Levels of D-ribose in urine were measured at different time intervals panel A. D-ribose levels in serum panel B and the brain panel C were determined within 3 days after dissection.

The expression and activity levels of ribokinase, transketolase TKT , 5-phosphoribosyl 1-pyrophosphate PRPP and glucose-6 phosphate dehydrogenase G6PD in the brain were measured with ELISA kits panel D and E. All values are expressed as the mean ± S.

TKT is a key enzyme in the nonoxidative branch of the pentose phosphate pathway PPP that is involved in the metabolism of D-ribose derivatives [ 33 , 34 ]. To investigate the mechanism of the D-ribose metabolic disorder in T1DM, we measured TKT expression and activity by ELISA.

As shown in Figure 1D and 1E , the expression and activity level of TKT were decreased remarkably in T1DM brain tissue. Other kinases, such as ribokinase, D-glucosephosphate dehydrogenase G6PD and ribose phosphate pyrophosphokinase PRPP , which also play roles in regulating D-ribose metabolism, did not show significant changes in expression or activity level in T1DM rats compared to control rats.

To demonstrate that TKT is linked to D-ribose dysmetabolism, we used BTMP to rescue the TKT change in T1DM rats since BTMP can increase the level of thiamine diphosphate and enhances TKT activity [ 35 ]. The results of the liver and kidney assays after BTMP treatment are shown in Supplementary Table 1.

As shown in Figure 2A , administration of BTMP increased brain TKT levels in both normal rats P P Figure 2B. Both the activity and expression Western blots of TKT in the liver and brain were significantly rescued after BTMP administration Supplementary Figure 2.

By contrast, D-ribose levels in both serum and brain were significantly decreased after BTMP administration Figure 2C , 2D. Under the experimental conditions, BTMP did not rescue or decrease FBG levels in T1DM rats Figure 2F. However, BTMP could partially rescue the body weights of T1DM rats but not the forepaw tension or insulin levels in the brain and serum Supplementary Figure 3.

That is, administration of BTMP can regulate the metabolism of D-ribose rather than D-glucose in rats via activation of TKT. Figure 2. Effect of benfotiamine BTMP on the levels of D-ribose, D-glucose and TKT in T1DM rats.

Conditions for the preparation of T1DM rats are shown in Figure 1. The expression levels of transketolase TKT in the brain panel A and liver panel B were measured with ELISA kits.

After 10 weeks of domestication, D-ribose levels in the serum panel C and brain panel D of rats were measured, and D-glucose levels were measured in the brain panel F. Fasting blood glucose FBG was measured every other week panel E.

According to McCrimmon and colleagues, both T1DM and T2DM are related to cognitive dysfunction [ 36 ]. Here, we investigated whether T1DM rats experienced cognitive impairment. First, compared with control rats, T1DM rats showed significantly fewer correct alterations in the Y maze test Supplementary Figure 4A.

In the Morris water maze test, the escape latency in the training session was significantly longer among T1DM rats Supplementary Figure 4B , and the percentage of time spent in the target quadrant in the probe trial was markedly lower for T1DM rats Supplementary Figure 4C.

T1DM rats also showed fewer platform crossings than did control rats, but the difference was nonsignificant Supplementary Figure 4D. Representative images of the performance path of the rats are shown in Supplementary Figure 4E.

These results indicated that T1DM rats exhibited cognitive impairment, which was regarded as type 1 diabetic encephalopathy. In addition, rats with type 1 diabetic encephalopathy also showed anxiety behavior based on open field and elevated plus maze assays Supplementary Figure 5.

To demonstrate whether cognitive impairment in T1DM rats was linked to D-ribose dysmetabolism, we tested the cognitive ability of T1DM rats with BTMP gavage. In the Y maze test, BTMP-gavaged T1DM rats exhibited significantly more correct alterations than in T1DM rats without BTMP gavage Figure 3A.

In the Morris water maze test, T1DM rats gavaged with BTMP spent less time searching for the platform than did T1DM rats without BTMP gavage Figure 3B. After platform withdrawal, the time spent in the target quadrant and the number of platform crossings were significantly higher in the T1DM group gavaged with BTMP than in that without BTMP gavage Figure 3C , 3D.

These data suggested that the alleviation of cognitive impairment by treatment with BTMP is related to a decrease in D-ribose in STZ-induced T1DM rats.

Figure 3. Rescue of spatial learning and memory abilities in T1DM rats with BTMP. Animal groups and treatments were as described in Figure 2 except that rats were subjected to Y maze and Morris water maze tests.

The accuracy of Y maze alternation was detected panel A. The escape latency panel B , percentage of time spent in the target quadrant panel C and number of platform crossings panel D were recorded.

Representative images of the performance path are shown panel E. As hyperphosphorylated Tau and the resultant neurofibrillary tangles and AGE are closely related to cognitive impairment [ 37 , 38 ], and high dose D-ribose treatment resulted in AGE aggregation and Tau hyperphosphrylation [ 39 ].

We wondered whether cognition-impaired T1DM rats exhibit Tau hyperphosphorylation and AGE accumulation along with a decrease in D-ribose. At the same time, markedly high AGE levels were detected in both the cortex and hippocampus.

These data suggest that T1DM rats suffer from Tau hyperphosphorylation as well as AGE accumulation in the brain. To investigate whether D-ribose dysmetabolism is linked to Tau hyperphosphorylation and AGE accumulation, we gavaged T1DM rats with BTMP and measured Tau phosphorylation and AGE levels.

As shown in Supplementary Figure 7 , AGE in both the cortex and hippocampus in BTMP-gavaged T1DM rats were significantly decreased compared with those without BTMP gavage. Furthermore, the results from the glycated serum protein GSP assay showed a marked decrease in GSP in T1DM rats after treatment with BTMP Supplementary Figure 7C.

These data demonstrated that D-ribose plays a role in AGE accumulation in T1DM rats. Along with the distinct decrease in AGE accumulation, Tau hyperphosphorylation was also reduced by BTMP administration. Tau phosphorylation levels AT8 and pSer in the cortex and hippocampus were significantly reduced, while nonphosphorylated Tau levels Tau-1 were increased in T1DM rats treated with BTMP compared with control rats Supplementary Figure 7.

That is, Tau hyperphosphorylation is related to the D-ribose dysmetabolism in T1DM rats. Neuronal loss is regarded as the most important pathological feature of age-related cognitive impairment.

We performed immunochemical experiments to determine neuronal death in the brains of T1DM rats. However, neuronal death in the brains of T1DM rats was greatly reduced by treatment with BTMP Figure 4A , 4B.

The decrease in D-ribose induced by BTMP occurred with the amelioration of pathological features, such as AGE accumulation, Tau hyperphosphorylation and neuronal death.

These data suggested that D-ribose dysmetabolism is closely related to cognitive impairment and that BTMP can be used as a potential medicine to rescue cognitive impairment via a decrease in D-ribose levels.

Additionally, the anxious behavior was observably improved after BTMP administration in the open field and elevated plus maze assays Supplementary Figure 8. Figure 4. Nissl staining of hippocampal neurons of rats treated with BTMP.

Animal groups and treatment were as described in Figure 2 except that hippocampal slices were prepared and stained with cresyl violet panel A. Numbers of necrotic neurons were counted under a microscope as described in the Materials and Methods panel B.

To detect the D-ribose level in T1DM patients, twenty-four participants 8 T1DM patients and 16 age-matched participants without diabetes mellitus were recruited for collection of fasting blood and morning urine. The summarized characteristics of the participants are shown in Supplementary Table 2.

We measured the concentrations of D-ribose in urine and serum by high-performance liquid chromatography HPLC. Figure 5. Comparison of D-ribose levels between T1DM patients and normal participants. D-ribose levels in serum panel A and urine panel B were measured by HPLC as previously described [ 17 ].

All values are shown as the mean ± S. T1DM results in long-term complications in the central nervous system, causing brain cellular dysfunction and cognitive deficits [ 40 ]. As reported in a recent study, T2DM patients suffer from D-ribose and D-glucose dysmetabolism [ 16 ].

In the present study, in addition to D-glucose, endogenous D-ribose was markedly increased in T1DM patients and STZ-induced T1DM rats, which also exhibited AGE accumulation and cognitive impairment accompanied with Tau hyperphosphorylation and neuronal death. Administration of BTMP decreased D-ribose levels and AGE accumulation and improved cognitive ability in T1DM rats.

BTMP also reduced Tau hyperphosphorylation, and neuronal death. All these data indicated that D-ribose dysmetabolism is associated with cognitive impairment in T1DM rat and that the administration of BTMP can ameliorate the loss of neurons and cognitive impairment via regulation of D-ribose.

As a very active aldose, D-ribose exists in urine [ 16 ], serum [ 41 ] and cerebrospinal fluid 0. AGE accumulation is one of the most important features in diabetes and its complications due to the high sugars levels [ 43 ]. The Panel considers that the effects observed in a subchronic toxicity study in Wistar rats could be the consequence of nutritional imbalances but that toxicological effects could not be ruled out [ 25 ].

In accordance with the Guidance for Industry, Center for Drug Evaluation and Research, U. Administration of an approximate dose 3. D-ribose can also give rise to Tau hyperphosphorylation and Aβ-like deposition in brain tissue and cause endoplasmic reticulum stress, which is toxic to cells and results in apoptosis [ 27 , 39 , 45 ].

Administration of D-ribose can increase hepatic triglyceride and water intake and decrease body weight in SD rats [ 24 ]. Furthermore, formaldehyde was regarded as a risk factor for age-related cognitive impairment [ 46 ]. These finding suggest that high dose D-ribose intake induces toxicity.

A high level of D-ribose was also found in the urine of T2DM patients, suggesting the dysmetabolism of D-ribose in T2DM [ 16 ]. Therefore, speculation that D-ribose dysmetabolism is involved in type 1 diabetic encephalopathy and its pathogenesis is reasonable.

T1DM rats showed a low level of TKT in brain tissue accompanied by a high level of D-ribose and pathological features of diabetic encephalopathy. Because D-ribose can be converted from D-glucose through the PPP [ 48 , 49 ], we measured G6PD and ribokinase.

However, G6PD and ribokinase did not show a marked change in T1DM rats except for TKT. These results indicated that STZ-induced T1DM rats did not suffer from dysfunction in G6PD, a key enzyme in the D-glycolytic pathway.

Many studies have shown that G6PD is upregulated [ 50 ] or downregulated [ 51 ] in STZ-induced DM rats. Epel demonstrated that the level of G6PD can be regulated by NADP, the critical factor in oxidative stress [ 52 , 53 ].

Many studies have also shown that the expression of G6PD regulates the generation of NADPH to alleviate oxidative stress [ 54 ], suggesting that G6PD would induce dynamic changes in DM.

Mendez et al. showed that neurons maintain the oxidation of G6PD through the PPP to sustain their antioxidant status [ 55 ]. In DM, apart from G6PD, TKT is also an important shunt key enzyme in the PPP [ 34 ]; thus, it makes sense that supplementation with BTMP, acting as a TKT activator [ 56 ], causes a reduction in oxidative stress and affects several anabolic reactions, which might also reduce the level of AGEs [ 57 , 58 ].

BTMP regulates the level of TKT that is directly involved in D-ribose metabolism [ 59 , 60 ]. In this studies, BTMP treatment upregulated TKT, decreased D-ribose levels and simultaneously rescued T1DM rats from diabetic-related encephalopathy but did not decrease D-glucose levels.

These data indicated that dysmetabolism of D-ribose with a decline in TKT function was involved in cognitive impairment in T1DM rats under the experimental conditions. Other laboratories also observed dynamic changes of TKT activity in different tissues in diabetes [ 61 , 62 ]. Consequently, TKT may be used as a potential drug target in the treatment of diabetic-related encephalopathy with high D-ribose levels.

BTMP gives a relatively wide range of actions on a number of cellular targets [ 58 ] such as treatment of inflammatory [ 63 ], peritoneal dialysis [ 64 ] and Tauopathy [ 65 ]. BTMP also plays a role in the metabolism of D-glucose [ 66 ]. According to Hammes and colleagues, BTMP activates TKT and prevents the activation of multiple pathways of hyperglycemic damage, such as the hexosamine pathway, the AGE formation pathway and the diacylglycerol-protein kinase C pathway, in diabetic animals [ 56 ].

Though activation of TKT through BTMP is downstream of the D-ribose pathway, which may not be direct evidence, the current work at least showed that a decrease in D-ribose levels could help the amelioration of cognitive impairment.

In fact, BTMP is closely related to D-ribose metabolism. BTMP markedly ameliorates the impaired spatial cognitive ability of T1DM rats in the Y maze and Morris water maze. Administration of BTMP decreases D-ribose levels in the brain, blood and urine of T1DM rats.

Chen and coworkers have indicated the correlation between D-ribose and the administration of BTMP in a ZDF rat animal model for diabetes [ 17 ]. Currently, a clinical trial on the treatment of cognitive impairment in Alzheimer's disease a pilot study with BTMP has been started and performed by Gibson and Jordan in the Burke Neurological Institute clinicaltrials.

Here, we would like to suggest that changes in D-ribose in blood and urine should be monitored and analysed in their clinical trials because BTMP can reduce D-ribose levels and ameliorate cognitive impairment in T1DM rats.

Neuronal loss may deteriorate cognitive ability. CA4 is a subfield of the hippocampus that is adjacent to the DG subfield [ 71 , 72 ]. The DG region, which is involved in long-term potentiation LTP and long-term memory, is associated with cognitive ability [ 73 , 74 ]. Here, we also found that T1DM rats showed anxiety-like behaviour Supplementary Figure 8.

In previous studies, BTMP was also shown to counteract anxiety-like behaviour [ 77 , 78 ]. The current work suggests that dysregulated D-ribose acts as a novel metabolite in cognitive impairment in T1DM rats by triggering protein glycation, Tau hyperphosphorylation and neuronal loss.

This viewpoint is based on the following observations. First, T1DM rats demonstrated high levels of D-ribose in the serum, urine and brain.

Second, T1DM patients also showed high levels of D-ribose in the urine and serum. Third, the expression and activity levels of TKT in the brain and liver of T1DM rats were reduced, which affected D-ribose metabolism [ 60 ]. Fourth, gavage of BTMP as the activator upregulated the expression of TKT in the brain and liver and decreased the levels of D-ribose but not those of D-glucose.

Fifth, administration of BTMP suppressed D-ribose levels and rescued cognitive impairment in T1DM rats in both the Y maze and Morris water maze assays, which confirmed the influence of D-ribose dysmetabolism.

Sixth, in T1DM rats with cognitive impairment, AGE accumulation and Tau hyperphosphorylation in the hippocampus and cortex were closely related to D-ribose dysmetabolism.

Finally, on the basis of previous work in this laboratory, AGE accumulation, Tau hyperphosphorylation and cognitive impairment were observed in a D-ribose-induced mouse model [ 27 , 79 ]. As described by other studies, both AGE [ 80 ] and Tau hyperphosphorylation [ 81 ] are associated with neuronal death [ 82 ] or loss [ 83 , 84 ], which can cause hippocampal atrophy [ 85 ] and result in cognitive impairment [ 86 , 87 ].

Therefore, D-ribose-induced neuronal loss may be an important contributor to cognitive impairment in T1DM rats. In conclusion, STZ-induced T1DM rats had high levels of D-ribose in their brain, serum and urine in addition to D-glucose.

TKT controlled D-ribose metabolism, and activation of TKT with BTMP decreased D-ribose levels, followed by a reduction in AGE formation, Tau hyperphosphorylation, neuronal death and cognitive impairment.

Thus, dysmetabolism of D-ribose is considered a novel pathological features in rats with T1DM and its complications. T1DM patients also show high levels of D-ribose. However, further investigations should be conducted on T1DM pathologies and complications related to D-ribose.

Male SD rats ~8 weeks, weighing ~ g were provided by Vital River Laboratory Animal Technology Co. The animals were housed in plastic cages measuring 45×30×26 cm 4 rats in each cage.

Rats were maintained under standard laboratory conditions, i. The handling of rats and experimental procedures were approved by the Animal Welfare and Research Ethics Committee of the Institute of Biophysics, Chinese Academy of Sciences Permit Number: SYXK Diabetes was diagnosed when the FBG level of rats was higher than Behavioral tests were carried out, and rats were sacrificed in the 10 th week.

Rats in the control group received a single intraperitoneal injection of citrate buffer and daily CMC gavage. Rats in the BTMP group received a single intraperitoneal injection of saline solution and daily BTMP gavage. Rats in the T1DM group received a single intraperitoneal injection of STZ and daily CMC gavage.

Behavioural tests were carried out, and rats were sacrificed in the 10 th week. The Y maze we used was composed of three equally spaced arms °; 47 cm long × 46 cm wide × 16 cm high, Beijing ZSdichuang Science and Technology Development Co.

In addition, hypoglycemia, a common complication of diabetes caused by low glucose levels in the blood, can lead to loss of energy for brain function and is linked to poor attention and cognitive function. Researchers are revealing more about how the brain and nervous system work — and translating those insights into new treatments.

Although the brain needs glucose, too much of this energy source can be a bad thing. A study in animals by researchers at the University of California at Los Angeles indicated a positive relationship between the consumption of fructose, another form of sugar, and the aging of cells, while a study, also using an animal model, conducted by a team of scientists at the University of Montreal and Boston College, linked excess glucose consumption to memory and cognitive deficiencies.

The effects of glucose and other forms of sugar on the brain may be the most profound in diabetes, a group of diseases in which high blood glucose levels persist over a prolonged period of time. Type 1 diabetes is a disease in which the immune system destroys the cells in the pancreas that produce insulin, a hormone used by the body to keep blood glucose levels in check.

Type 2 diabetes, caused by dietary and other environmental factors, is a condition in which cells become overwhelmed by insulin and fail to properly respond; they become resistant to insulin.

Vera Novak, MD, PhD. Long-term diabetes—either type 1 or type 2—has many consequences for the brain and for neurons in the brain, says Novak. It can cause the brain to atrophy or shrink. And it can lead to small-vessel disease, which restricts blood flow in the brain, causing cognitive difficulties and, if severe enough, spurring the development of vascular dementia.

In her laboratory, Novak is studying ways to prevent these effects in people with type 2 diabetes. One of these ways involves a nasal spray called intranasal insulin INI.

Citation: Han C, Lu Y, Wei Y, Liu Y, He R D-Ribose Induces Cellular Protein Glycation and Impairs Mouse Spatial Cognition. PLoS ONE 6 9 : e Received: July 28, ; Accepted: August 14, ; Published: September 8, Copyright: © Han et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by grants from the Project CB; CB , the Natural Scientific Foundation of China NSFC , , CAS-KSCX2-YW-R, KSCX2-YW-R, QCAS Biotechnology Fund GJHZ and by Beijing Natural Science Foundation Ribosylation of intracellular protein and the mechanism of cell death.

The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing interests: The authors have declared that no competing interests exist. Non-enzymatic glycation of proteins by reducing saccharides such as D-glucose Glc and D-ribose Rib is a post-translational modification process [1] , leading to the formation of fructosamine [2] and advanced glycation end products AGEs [3].

The role of Glc in the glycation of proteins has been widely studied, however the role of other reducing monosaccharides such as Rib in glycation and their resulting effects on cell metabolism has received much less attention. D-ribose is a naturally occurring pentose monosaccharide present in all living cells including the blood and is a key component of many important biomolecules such as riboflavin i.

As a reducing monosaccharide, Rib has the ability to react with proteins to produce glycated derivatives. Glycation with Rib ribosylation gives rise to AGEs more rapidly than glycation with Glc which requires a relatively long time [7].

Rib, however, is also closely associated with many fundamental processes in cellular metabolism. For this reason, glycation of proteins with Rib needs to be addressed and investigated. The rate of glycation depends upon monosaccharide concentration and anomerization rate and is inversely proportional to the number of carbon atoms in the reducing monosaccharide [8].

Under physiological conditions, the anomerization rate of Rib is much higher than that of Glc. This unstable aldofuranose ring is vulnerable to reactions with amino groups, giving rise to its high efficiency in protein glycation.

Therefore, comparing ribosylation with glucosylation should provide new clues for clarifying some of the important complications caused by advanced glycation end products in vivo. In vitro studies on the role of Rib in glycation have been carried out.

Rib can glycate rat tail tendon collagen in vitro and the structure of the collagen is significantly altered by Rib-induced glycation [10]. Luciano and colleagues prepared glycated fetal calf serum with Rib and found that while ribosylation reduces the proliferation of pancreatic islet beta-cells, cell necrosis and cell apoptosis rate increase correspondingly [11].

Ribosylated bovine serum albumin polymerizes and forms globule-like aggregates with high cytotoxicity [12]. However, the relationship between ribosylation and neurodegenerative diseases is still unknown.

In this laboratory we have observed that glycation induces inactivation and conformational change of D-glyceraldehydephosphate dehydrogenase [13] , [14]. We have also compared the characteristics of ribosylation on neuronal Tau protein [15] , and α-synuclein [16] with those of glucosylation in vitro , showing that ribosylation occurs much more rapidly than glucosylation.

Ribosylated neuronal protein is much more cytotoxic than glucosylated protein. Nevertheless, little is known about whether Rib can rapidly induce glycation in cells and the AGEs produced by ribosylation can impair the cognitive function. Here, we treated cultured cells and mice with Rib and Glc to compare ribosylation and glucosylation for the production of AGEs.

We found that Rib reacted rapidly with proteins and produced significant amounts of AGEs in cultured cells and mouse brain tissues, and that accumulation of AGEs impaired mouse spatial cognition. This finding implies that Rib-derived AGEs may be related to impairments of learning and memory ability.

To investigate whether Rib leads to decreases in cell viability, SH-SY5Y human neuroblastoma SH-SY5Y cells and Human embryonic kidney T HEKT cells were incubated with D-ribose or D-glucose at different concentrations.

Cell viability was measured by MTT assays at 2 and 3 days after addition of the monosaccharide. MTT assays also gave the same results after 3 days of Rib treatment Fig 1H and J. Furthermore, the number of SH-SY5Y and HEK cells was markedly lower after treatment with 50 mM Rib for 3 days compared with Glc-treated and control cells Fig 1A to F.

The morphology of SH-SY5Y cells was observed by inverted contrast microscopy after incubation with 50 mM Rib A , or 50 mM Glc B for 3 days. Untreated cells were used as controls C. HEKT cells treated with the same concentration of Rib D , Glc E and control cells F were imaged under the same conditions.

SH-SY5Y G and H and HEKT I and J cells were incubated with Rib or Glc as indicated and cell viability was measured using the MTT assay at day 2 G and I , and day 3 H and J after addition of the monosaccharides. It is known that AGEs have cytotoxicity [15] , [17] and can inhibit cell proliferation [18].

Rib and Glc react with protein amino groups to initiate a non-enzymatic glycation process which results in AGE formation. Thus, we detected the presence of AGEs in SH-SY5Y, HEK cell lines and primary cultured hippocampal neurons incubated with Rib for 2 days by Western blotting.

However, in the presence of Glc, the level of AGEs did not increase significantly under the experimental conditions used. Similarly, the level of AGEs in both HEKT cells and primary cultured hippocampal neurons was also enhanced significantly after Rib treatment Fig 2B and C.

These results indicate that Rib is much more active in protein glycation resulting in high yields of AGEs and reduced cell viability. SH-SY5Y cells panel A , HEKT cells panel B and primary cultured rat neurons panel C were treated with Rib and Glc as indicated for 2 days.

AGEs were detected with anti-AGEs 6D12 monoclonal antibody. β-Actin was used as a loading control. The control value was set as 1. All values are expressed as means ± S.

Having determined that Rib but not Glc is able to glycate proteins rapidly and produce high levels of AGEs in cultured cells in vitro , we investigated whether Rib is able to induce AGE formation in vivo.

Mice were injected i. with Rib as indicated for 30 days and serum was taken for assays of glycated serum protein. Mice injected with Glc and saline were used as controls. Having established that injection of Rib leads to an increase in glycated serum proteins, we measured changes in serum AGE formation in mice treated with Rib and Glc to determine whether high levels of glycation lead to AGE production Fig 4.

Strikingly, serum AGEs were markedly elevated in the sera of mice that had been injected with Rib. Those treated with Glc were not significantly different from the control group. Similar results were also obtained when the anti-pentosidine antibody was used. Serum pentosidine level was markedly increased in the presence of Rib both 0.

These results demonstrate that Rib significantly elevates the glycation of proteins in the blood resulting in accelerated AGE formation under our experimental conditions.

Conditions for the injection of monosaccharides were the same as those given in Fig 3 , except that serum AGEs were detected with an anti-AGEs monoclonal antibody panel A and an anti-pentosidine monoclonal antibody panel B.

Serum albumin level was used as a loading control. The saline control value was set as 1. Rib can pass through the blood-brain barrier and enter the brain by simple diffusion [19].

To investigate whether injected Rib can elevate the glycation of proteins in the brain, we measured AGEs in the mouse brain by Western blotting. As shown in Fig 5 , intraperitoneal injection of Rib led to the formation of significantly more AGEs in the mouse brain. However, Glc did not have a significant effect on AGE formation in the brain compared to the control.

This suggests that Rib can react effectively with proteins and increase AGEs in the mouse brain. Conditions for the injection of Rib were the same as those given in Fig 3 , except that AGEs in the mouse brain were detected by Western blotting using anti-AGEs 6D12 monoclonal antibody, panel A.

β-Actin level was used as a loading control. Quantification results are shown in panel B. To confirm the effect of Rib on accelerating the formation and accumulation of AGEs in the brain, we performed immunohistochemistry staining on microtome sections of the mouse brain Fig 6.

Compared with the control group, AGEs were observed to increase throughout the hippocampus of mice that had been injected with Rib for 30 days. However, no obvious differences in the hippocampus were found in the Glc-treated and control mice groups.

Furthermore, AGE signals were more clearly evident in the cortex of Rib-treated mice, compared with those treated with Glc. This indicates that the rapid formation of Rib-induced AGEs occurred in both the hippocampus and cortex.

AGEs in the mouse brain were detected by immunohistochemistry using anti-AGEs monoclonal antibody. We used immunofluorescent staining to further demonstrate that Rib is able to induce AGE formation in the mouse brain. As shown in Fig 7 , AGE signals were clearly visible in the cortex of mice treated with Rib but not in those treated with Glc or saline.

The fluorescent signals of AGEs were mainly localized outside the nucleus. Similar results were also observed in the hippocampus of the Rib group though the signals were relatively lower than those in the cortex.

AGEs in the mouse brain were detected by immunofluorescent staining. The brain sections were double-labeled for AGEs green and nuclei blue. AGEs, which have been found in the brains of senile dementia patients [20] , are cytotoxic [12] , [17]. To assess changes in the spatial learning and memory of mice whose brain AGE levels were elevated after injection of Rib, we tested their behavior in the Morris water maze.

During the training session, all mice improved their performance as indicated by shortened escape latencies over successive days, and mice from each treatment group had the same level of performance no significant individual effect was observed in the first three trials on day 1 prior to treatment.

Escape latencies of mice injected with Rib 0. Conditions for the injection of Rib were the same as those given in Fig 3. The length of time mice took to find the hidden platform was recorded as latency of escape during each of the seven training days panel A and B.

The length of the searching time spent in the quadrant when the platform was removed during the probe trial is shown in panel C. Withdrawal of the platform induced a general tendency to swim in the quadrant where the platform was previously located and in the platform zone, in preference to other equivalent zones.

Control, Rib 0. These results indicate that spatial learning and memory ability in Rib-treated mice are significantly impaired. As a reducing saccharide, Rib reacts with protein amino groups to initiate a post-translational modification process widely known as non-enzymatic glycation [1].

This reaction proceeds from reversible Schiff bases to stable, covalently-bonded Amadori rearrangement products. Once formed, the Amadori products undergo further chemical rearrangements to form irreversibly bound AGEs, which are a heterogeneous group of structures including pyrraline, pentosidine, crossline, and carboxymethyl-lysine [3].

As described previously, Rib is much more active in protein glycation than Glc in vitro [12] , [15] , [16]. Here, we also found that Rib reacts rapidly with proteins and showed that Rib treatment results in a significantly higher level of AGEs both in cultured cells and in the mouse serum and brain.

This demonstrates that AGEs result from ribosylation not only in mixtures of Rib and proteins in a test-tube, but also in cultured cells, and in the mammalian serum and brain.

Even though glycation of proteins with reducing saccharides has been widely studied, the formation of monosaccharide-induced intracellular AGEs has not previously been observed. Here, 10 mM Rib enhanced AGE formation in cultured cells, and diminished cell viability.

This work is the first to show that Rib enhances the yield of AGEs in HEKT and SH-SY5Y cells and primary cultured hippocampal neurons.

Rib showed significantly higher cytotoxicity than Glc in cell culture. The high cytotoxicity of Rib may result from the rapid formation of AGEs as a result of ribosylation under these experimental conditions.

Furthermore, this monosaccharide also enhances the yield of AGEs in both the hippocampus and cortex of the mouse brain after intraperitoneal injection. Impairment of spatial cognition was observed to be coincident with these increases in intracellular AGEs when Rib-treated mice were tested in the Morris water maze.

Glc, however, was unable to elevate the yield of AGEs under our experimental conditions. This clearly demonstrates that an overload of Rib may result in a high level of AGEs in the brain and neurons. We would like to emphasize that Rib is much more effective than Glc in the glycation of proteins not only in vitro, but also in vivo.

The intracellular AGE-enhanced cell model and the AGE-enhanced cognitive impairment mouse model successfully established here can be used for further investigation of the mechanisms behind the phenomena observed. As shown in the results of Western blotting Fig 2 and 5 , a low level of glycated proteins or AGEs is already present under physiological conditions in both cells and the mouse brain since cells and blood contain certain concentrations of reducing saccharides.

High levels of Rib are not only able to enhance AGE formation in vivo , but also induce dysfunction of spatial cognition. This is the first time that Rib-induced cognitive impairment has been observed in mice.

The spatial learning and memory ability of mice markedly declined after 30 days of Rib administration. However, those that were injected with the same concentration of Glc did not show significant impairments of spatial cognition compared with saline controls.

D-Ribose is an essential monosaccharide with a pivotal role in our metabolism and energy production 2 Jinni Hong et al, D-ribose induces nephropathy through RAGE-dependent NF-κB inflammation Arch Pharm Res.

Several research endeavors suggest that D-Ribose could benefit heart health 4 Shuai Li et al, D-ribose: Potential clinical applications in congestive heart failure and diabetes, and its complications Review Exp Ther Med.

It may help improve the energy metabolism in the heart, offering a quality of life boost for those with specific heart conditions 5 Shuai Li et al, D-ribose: Potential clinical applications in congestive heart failure and diabetes, and its complications Review Exp Ther Med.

But a word of caution here: Always seek professional healthcare advice before beginning any new supplementation, especially if you have an existing heart condition. This sugary molecule could improve cognitive performance and mood 7 Jacob E Teitelbaum et al, The use of D-ribose in chronic fatigue syndrome and fibromyalgia: a pilot study J Altern Complement Med.

D-Ribose also plays a role in the pentose phosphate pathway, producing NADPH 8 Guoyao Wu et al, Glutathione metabolism and its implications for health J Nutr. Moreover, D-Ribose assists in nucleotide synthesis.

These nucleotides are the building blocks for our DNA and RNA — essential for cell growth, repair, and communication 9 Diane E. Mahoney et al, Understanding D-Ribose and Mitochondrial Function Adv Biosci Clin Med. Intriguingly, a cross-sectional study on patients aged 60 and older associated elevated urinary D-Ribose levels with increased cognitive decline rates 10 Xinyi Zhu et al, Urine D-ribose levels correlate with cognitive function in community-dwelling older adults BMC Geriatrics.

This could mean D-Ribose might be a practical, non-invasive biomarker for mental impairment progression. However, no negative correlation between D-Ribose and cognitive function was seen in episodic memory, working memory, or processing speed tests.

A systematic review of animal studies reveals that D-ribose treatment might induce cognitive impairment, with more significant effects at higher doses 13 Ying Song et al, A systematic review and meta-analysis of cognitive and behavioral tests in rodents treated with different doses of D-ribose Front.

Aging Neurosci. In rodents, D-ribose affected spatial learning tasks but not spatial memory tasks. Moreover, high D-ribose doses increased AGEs in the brain and serum, potentially causing cognitive impairment.

D-Ribose is a monosaccharide like glucose, but its effect on blood glucose might be more adverse than we thought. Interestingly, D-Ribose outperforms other sugars in producing glycated serum protein GSP , a precursor to chronic diabetes complications 15 Yao Chen et al, d-Ribose contributes to the glycation of serum protein Biochim Biophys Acta Mol Basis Dis.

A notable observation here is that D-Ribose interacts more rapidly than glucose with non-enzymatic proteins in diabetic patients, leading to the formation of AGEs. The binding of these AGEs to their receptors triggers a series of events: the upregulation of vascular growth factors, increased vascular permeability, angiogenesis, and local inflammation 16 Alejandra Planas et al, Advanced Glycations End Products in the Skin as Biomarkers of Cardiovascular Risk in Type 2 Diabetes Int J Mol Sci.

This AGE-receptor pathway activation also stimulates oxidative stress reactions, contributing to chronic diabetes complications. Although D-Ribose might temporarily reduce blood sugar levels, its long-term accumulation can worsen diabetes complications.

Consequently, monitoring D-ribose levels and glycosylated proteins in diabetic patients might help forecast potential clinical complications, providing an opportunity for proactive prevention. Given the potential brain-health complications linked to high D-Ribose levels, classifying D-Ribose as a nootropic might be a stretch.

However, numerous natural nootropics can prevent cognitive decline and boost brain health. As you can see, the question as to whether you can benefit from D-ribose supplementation is tricky.

While there are many positive aspects, many adverse outcomes correlate to high serum d-ribose. As more research comes to light, we can hopefully get a more straightforward answer, especially regarding brain health.

Check with your doctor to determine whether D-ribose is right for you! Join us at Holistic Nootropics as we journey through all facets of health and wellness. Save my name, email, and website in this browser for the next time I comment. Keep in mind that we may receive commissions when you click our links and make purchases.

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However, the relationship between variations in RIB levels and depression as well as potential RIB participation in depressive disorder is yet unknown. Here, a reanalysis of metabonomics data from depressed patients and depression model rats is performed to clarify whether the increased RIB level is positively correlated with the severity of depression.

The results show that RIB caused intestinal epithelial barrier impairment and microbiota-gut-brain axis dysbiosis. These microbial and metabolic modules are consistently enriched in peripheral fecal, colon wall, and serum and central hippocampus glycerophospholipid metabolism.

These findings suggest that the disturbances of gut microbiota by RIB may contribute to the onset of depressive-like behaviors via regulating glycerophospholipid metabolism, and providing new insight for understanding the function of microbiota-gut-brain axis in depression.

D-ribose RIB is a naturally occurring monosaccharide that is found in riboflavin-containing foods such as wheat bran, eggs, and meat. Meanwhile, because RIB can bypass part of the pentose pathway to produce d -ribosephosphate for the production of energy, it has been utilized as a daily nutritional or energy supplement 1 , notably for patients with chronic fatigue syndrome and coronary artery disease 2 , 3.

RIB is also a crucial component of several important biomolecules including adenosine and adenosine triphosphate, which are involved in a variety of metabolic activities 4.

However, as described by the European Food Safety Authority, the toxicological effects of RIB should not be ignored 5. Several studies have reported that RIB can be involved in the onset of encephalopathy 6 , 7. Depression is one of the most prevalent serious mental disorders, characterized by a lack of interest, pessimism, appetite loss, and even suicidal behavior 8 , 9.

Finding relevant risk variables is crucial for depression prevention and screening. Given that RIB has not been widely reported in depressive disorder and that a high-sugar diet may be an environmental risk factor for depression 11 , 12 , we recently gave normal mice prolonged RIB supplementation and found that these mice exhibited depressive-like behaviors and histological alterations, including obviously condensed and deeply-stained pyramidal cells in the hippocampus This finding implies that RIB has a significant impact on the development of depression.

The key scientific concerns that we will research, however, are whether the variation in RIB level was associated with depression and the underlying biological mechanism of RIB implicated in depressive illness. Of note, some other monosaccharides, such as fructose and glucose, have been linked to changes in the gut microbiota, leading to microbial metabolite disorder in rodents 14 , A high intake of sugar can cause enteric dysbacteriosis.

The latter increases the permeability of the intestinal mucosa, and results in abnormalities in intestinal immunity and glucolipid metabolism Moreover, hyperglycemia would increase the permeability of the intestinal barrier, giving microbes a better chance to enter the body and causing the proliferation of pathogenic bacteria Our groups previously found that depression was linked to altered gut microbiomes 19 , and germ-free mice exhibit depressive-like behaviors after receiving gut microbiota from depressed patients 19 , Furthermore, a recent study demonstrates that exogenous RIB can affect gut microbial architecture As a result, we proposed that changes in microbiota might account for the connection between RIB intake and depression.

At first, we reanalyzed the metabonomics data from our earlier studies to clarify the change of RIB in the urine of depressed patients and in the hippocampus of depression model rats. Then, in the current study, to further investigate the possible mechanism of RIB-induced depression, eight weeks of RIB-fed mice were constructed.

The intestinal barrier impairment was evaluated using hematoxylin and eosin, immunohistochemistry, and electron microscopy. The distinct gut microbiota was initially identified by 16S rRNA gene sequencing analysis. Moreover, by systematic analysis of relevant biological samples, including peripheral fecal, colon wall, and serum and central hippocampus specimens from the RIB-fed mice and control mice, comparative untargeted metabolomics was used to capture the functions of the altered gut microbiome.

Finally, by integrating these multi-omics data, we sought to understand how the gut microbiota contributed to the development of depressive-like behaviors and to pinpoint a putative way between the gut and the brain in RIB-fed mice. The results indicate that a tight connection may exist between elevated RIB and depressive illness.

c There was a negative relationship between the percentage of sucrose preference SP and the relative abundance of RIB. d There was a significant positive correlation between immobility time IT and the relative abundance of RIB.

Data are the means ± standard error of mean. These results further confirm that RIB-fed induced depressive-like behaviors in mice. a , b Both body weight a and fasting blood b were similar between the two groups. d Total distance was similar between the CON and RIB groups.

e Center distance was significantly lower in the RIB group. f Compared to the CON group, the RIB group had a significantly higher immobility time. Homeostasis in the gut is important for brain function.

We investigated whether RIB feeding perturbed colonic homeostasis, including colonic barrier and gut microbiota. Electron microscopy also showed severe mitochondrial swelling, injury of tight junction and gap junction domains, reduced numbers of the desmosome, and increased distance between adjacent epithelial cells in the colon of RIB-fed mice Fig.

Moreover, immunohistochemistry analysis indicated that the expression of Occludin Fig. M mitochondria, TJ tight junction, De desmosome. Next, gut microbiota diversity and composition in response to RIB were analyzed using 16S rRNA gene sequencing.

There were no significant differences in alpha diversity between the two groups Supplementary Fig. As shown in Fig. The relative abundances of each phylum were described in Supplementary Data 1. a Principal coordinate analysis model showed that there were significantly differential gut microbiota compositions between CON and RIB-fed mice.

b Firmicutes, Bacteroidetes, and Verrucomicrobiota were the three major bacterial phyla in both groups. c In total, 22 differential genera responsible for the discrimination between CON and RIB-fed mice were identified using linear discriminant analysis effective size.

The relative abundance of glycerophospholipid metabolism was highest among these pathways. A1: Phenylalanine, tyrosine, and tryptophan biosynthesis; A2: Lysine degradation; A3: Valine, leucine, and isoleucine biosynthesis; A4: Histidine metabolism; L1: Secondary bile acid biosynthesis; L2: Glycerophospholipid metabolism; L3: Arachidonic acid metabolism; L4: Primary bile acid biosynthesis; L5: Fatty acid elongation; L6: Fatty acid biosynthesis.

CD center distance, IT immobility time, SP sucrose preference. Using linear discriminant analysis effective size, 22 differential genera responsible for the discrimination between CON and RIB-fed mice were identified Fig.

Kyoto Encyclopedia of Genes and Genomes KEGG pathway analysis Fig. The results Fig. In total, there were metabolites successfully annotated Supplementary Data 3. The built orthogonal partial least-squares discriminant analysis OPLS-DA model using microbial metabolites in feces showed that the RIB-fed mice were separate from CON with no overlap, suggesting the divergent microbial metabolic phenotypes between the two groups Fig.

The results of permutation testing demonstrated that this model was valid and not over-fitting Supplementary Fig. Detailed information on these microbial metabolites was described in Supplementary Data 4. The heat map consisting of these differential microbial metabolites showed a consistent clustering pattern within the individual groups Fig.

a OPLS-DA model showed that the two groups had significantly different fecal metabolic phenotypes. c KEGG pathway classification showed that these metabolites were mainly annotated into the metabolism category. d Using online software MetaboAnalyst, four significantly dysregulated metabolic pathways in KEGG metabolism category classifications at level 3 were identified via hypergeometric test each dot represents a KEGG path, the dot size represents the impact value, and the dot color represents the p-value, the more important the differential metabolites were in this pathway, the larger the dot.

In addition, KEGG pathway classification showed that these differential microbial metabolites were mainly annotated into the metabolism category Fig. Using online software MetaboAnalyst, four significantly affected metabolic pathways in KEGG metabolism category classifications at level 3 were identified via hypergeometric test Fig.

In total, metabolites were successfully annotated in the colon Supplementary Data 5. The built OPLS-DA model showed divergent metabolic phenotypes in the colon between the two groups Supplementary Fig. KEGG pathway classification showed that these differential microbial metabolites were mainly annotated into the metabolism category; and using the online software MetaboAnalyst, seven significantly affected metabolic pathways in KEGG metabolism category classifications at level 3 were identified via hypergeometric test Fig.

a Seven significantly dysregulated metabolic pathways were found using hypergeometric tests in colon tissue. b Eight significantly dysregulated metabolic pathways were identified using hypergeometric tests in blood. c Three significantly dysregulated metabolic pathways were identified using hypergeometric tests in hippocampus.

The online software MetaboAnalyst was used to conduct pathway analysis. Each dot represents a KEGG path, the dot size represents the impact value, and the dot color represents the p -value.

The more important the differential metabolites were in this pathway, the larger the dot. d A heat map representation comprising all the differential metabolites from the colon, blood, and hippocampus showed a consistent clustering pattern within the individual groups.

S3b and Supplementary Data 8. Using online software MetaboAnalyst, eight significantly affected metabolic pathways in KEGG metabolism category classifications at level 3 were identified via hypergeometric test Fig. S3c and Supplementary Data Using online software MetaboAnalyst, three significantly affected metabolic pathways in KEGG metabolism category classifications at level 3 were identified via hypergeometric test Fig.

A heat map representation comprising all the differential metabolites from colon, blood, and hippocampus showed a consistent clustering pattern within the individual groups Fig. This phenomenon was also observed in our previous nonhuman primate model of depression As such, the different components of a given metabolic pathway might synergistically modulate the function of the MGB axis in different tissues.

Thus, weighted correlation network analysis was used here to cluster the identified differential metabolites into the metabolic modules of the MGB axis.

The results showed that there were seven different modules, in which four modules blue, red, black, and turquoise were significantly correlated with at least one type of depressive-like behavior Fig.

SP was significantly correlated with three metabolic modules blue, red, and black , and IT was significantly correlated with the turquoise metabolic module. Red and green squares indicated positive and negative correlations, respectively. b The differential metabolites in the turquoise metabolic module were significantly correlated with IT mainly belonged to lipid metabolism, especially PE and PC.

c The differential metabolites in three metabolic modules blue, red, and black were significantly correlated with SP mainly belonging to PC, fatty acyls, organic compounds, and carboxylic acids and derivatives.

Differential metabolites belonging to lipid metabolism were marked using different colors except for the gray, and other differential metabolites were marked using gray circles. Circle size indicated the abundance of the metabolites belonging to this node.

BW body weight, CD center distance, CL cardiolipin, FB fasting blood, IT immobility time, PA phosphatidic acid, PC phosphatidylcholine, PE phosphatidylethanolamine, PI phosphatidylinositol, PS phosphatidylserine, SP sucrose preference, TD total distance.

Module-trait analysis showed that the differential metabolites in the turquoise metabolic module significantly correlated with immobility time were mainly involved in peripheral and central glycerophospholipid metabolism within the MGB axis Fig. Details regarding the module and chemical class of each compound were shown in Supplementary Data The abovementioned findings indicated that glycerophospholipid metabolism might play an important role in the crosstalk of gut microbiota and the brain.

The numbers associated with the metabolite names were just codes, for example, we used PC39 to represent 1-heptadecanoyl-glycerophosphate detailed information about the codes representing metabolites is shown in Supplementary Data PC phosphatidylcholine, PE phosphatidylethanolamine.

These results indicated that glycerophospholipid metabolism, especially PC and PE, might be the important bridge of gut microbiota in affecting brain functions. Dietary sugars, like fructose and glucose, are associated with psychosis-related higher brain dysfunctions 12 , Our previous study provided evidence that another simple sugar, RIB, could lead to depressive-like behaviors, and we demonstrated in mice that this was connected with altered hippocampus metabolic and transcriptome profiles 13 , but how the brain is affected by RIB remains poorly understood.

In this study, we clarified that the RIB level was significantly increased in the depressed patients and depression model rats, and there was an obvious correlation between the change of RIB and the severity of depression disorder.

However, these studies only involved untargeted metabolomics analysis, which provides relative metabolite abundance rather than absolute quantification The results further suggested that high levels of RIB were correlated with depression. We observed the RIB-fed mice were characterized by intestinal epithelial barrier impairment, alterations of microbial composition, function, and metabolic pathways of the MGB axis.

Meanwhile, the altered microbial and metabolic modules linked the gut microbiome with dysregulation of peripheral and hippocampus glycerophospholipid metabolism in RIB-fed mice. To our knowledge, this is the first report of RIB influencing gut microbiota, and gut dysbiosis may be responsible for mediating the depressive-like behaviors seen in RIB-fed mice by regulating the MGB metabolism.

We found that the RIB-fed mice had considerably impaired intestinal barrier as compared to the CON. The gut barrier function was regulated by the gut microbiota 24 , As a result, we deduced that the RIB might disrupt the gut microbiota, and that the gut dysbiosis would subsequently lead to depression via the MGB axis.

Akkermansia belonged to the Verrucomicrobiota phylum. Khan S et al. The mucus-degrading bacterium Akkermansia would regulate intestinal homeostasis and preserve the integrity of the gut barrier The increased Akkermansia might be the cause of intestinal barrier impairment in RIB-fed mice.

Interestingly, recent research has shown that Akkermansia has beneficial roles in human health 30 ; nevertheless, there is also strong evidence that Akkermansia promotes the etiology of colitis Similarly, Akkermansia was detected in much higher abundance in individuals with severe depressive symptoms, according to Zhang et al.

These contradictory findings might be the consequence of differences in participants, sequencing, and analytical approaches. Furthermore, we found that This result is in line with our earlier research that At the phylum level, disturbances of Firmicutes have been identified as a possible hallmark of depression 19 , Accordingly, these results suggested that RIB would induce the gut microbiota disordered, and that the bacterial phylum Firmicutes disturbances might be a significant contributing factor to RIB-caused depressive-like behaviors.

According to previous research 36 , RIB levels in human urine were positively correlated with serum triglyceride levels, and Sprague-Dawley rats given RIB injection had considerably higher hepatic triglyceride levels, suggesting that RIB might regulate lipid metabolism. Besides, lipids are crucial for brain neuronal activity 37 , and the lipid composition of the brain has a significant impact on emotional behavior and subjective mood Here, we found that both altered microbial and metabolic modules involving glycerophospholipid metabolism were highly correlated with depressive-like behaviors in RIB-fed mice.

Our previous studies revealed that the gut microbiota would significantly affect the glycerophospholipid metabolism in the mouse brain 41 , Meanwhile, the glycerophospholipid metabolism in the hippocampus was disturbed in the chronic unpredictable mild stress rat model of depression Thus, we concluded that in RIB-fed mice, peripheral and central glycerophospholipid metabolism was regulated by gut dysbiosis, which might be a contributing factor to the development of depressive-like behaviors.

In addition, we found that PC and PE had greater contributions to the overall correlation between differential genera and glycerophospholipids. Our previous study has also shown that they are remarkably increased in depressed patients and have favorable associations with depression severity Moreover, PC and PE are critical components of neuronal membranes.

The phospholipase A2 can deacylate PC and PE into lysophosphatidylcholine and lysophosphatidylethanolamine, which are then converted into glycerophosphocholine and glycerophosphoethanolamine. Patients with depression have increased amounts of the neurotransmitter acetylcholine, which is produced by glycerophosphocholine Higher levels of glycerophosphoethanolamine have been found in the white matter of depressed individuals In light of this, our findings suggested that in RIB-fed mice, PC and PE might act as a link between gut dysbiosis and depressive-like behaviors.

Several limitations of this study are listed as follows: First, it was conducted in mice; however, human gut microbiota compositions do not exactly match those of rodents, clinical trials are required to further confirm the reported effects of RIB on human gut microbiota composition and function.

Second, we only focused on one region of the brain associated with emotions the hippocampus , whereas chronic stress caused lipidomic changes in a region-specific manner, and the disturbances of lipid metabolism in the prefrontal cortex were more obvious than those in the hippocampus As a result, future studies should take other brain regions to uncover more novel clues on the interactions between gut microbiota and RIB-caused depression.

Third, our data do not completely rule out the possibility of other direct effects of RIB on the host, even though we thoroughly defined host peripheral and hippocampus metabolism implicated in RIB-induced gut dysbiosis. Future study on fecal microbiota transplantation is needed to clarify how the MGB axis links RIB to aberrant emotional-associated behavior.

Fourth, due to the dose of ribose being a constant here, we could not use mediation analyses to assess whether ribose directly induced depression or indirectly induced depression through influencing the gut microbiome or gut-brain axis glycerophospholipid metabolism.

Future studies should explore the role of RIB directly or indirectly in depression by designing gradient RIB dose experiments or modulating the important differential variables identified in this study bacterial taxa or metabolites.

Considering that RIB induced depressive-like behavior, which correlated with intestinal barrier damage and gut microbiota imbalance, the issue of whether intervention or reversal of intestinal barrier damage and gut microbiota imbalance could circumvent the effects of RIB requires further investigation in future studies.

Sixth, due to the limitations of technologies and funds, we did not conduct further experiments to validate the functional relationship of RIB with the glycerophospholipids metabolism pathway. In conclusion, to our knowledge, this is the first study to reveal that oral RIB results in depressive-like behaviors, which may be partially explained by changes in microbial composition, function, and metabolism of the MGB axis.

This study highlights that simple sugars like RIB can have adverse effects on gut microbiota, MGB axis metabolism, and mental health. The relative abundance of RIB was defined as RIB level in metabonomics.

The MDD patients were recruited from the Psychiatric Center of the First Affiliated Hospital of Chongqing Medical University, and the HC were recruited from the Medical Examination Center of the First Affiliated Hospital of Chongqing Medical University.

All included individuals were ethnically homogenous Han Chinese. The Hamilton Depression Scale score of MDD patients has to be more than 17, and all of them were first-episode drug-naïve MDD patients. The human data used and study protocol have received ethical approval from the Medical Research Ethics Committee of Chongqing Medical University.

All ethical regulations relevant to human research participants were followed, and a statement confirming that informed consent was obtained. Following the protocol 49 , a typical depression phenotype caused by chronic mild stress was modeled in rats using a chronic social defeat stress paradigm.

Animals were housed in a specific pathogen-free vivarium under standard conditions. All RIB group mice were administered the RIB 3. Louis, MO, USA in drinking water for eight weeks. The dosage and time were determined by previously published studies 13 , Thus, here we used 3.

The mice in the CON group were given access to normal purified water for feeding. Since food is one of the primary factors influencing changes in the gut microbiome, all the mice were fed the same food standard laboratory mouse diet to rule out any possible impacts.

To reduce any environmental effects, all mice were kept in the same room and habituated to the experimental setting for at least a week, and the drinking water added RIB was replaced once every two days.