Diabetic central neuropathy, which refers to neuron damage and may eventually result in dementia, is one of the most common complications of type 2 diabetes. Sitagliptin has been widely prescribed as pharmacotherapy for the treatment of patients with type 2 diabetes. However, the effects of sitagliptin on diabetic central neuropathy and intestinal ecosystem remain poorly studied. Our work is the first to consider that sitagliptin ameliorates diabetic central neuropathy via preventing the hippocampal neuronal damage and alters structure of gut microbiota in type 2 diabetic rats. Our current study demonstrates that sitagliptin might alleviate diabetic central neuropathy through triggering microbiota-gut-brain axis.
Diabetic central neuropathy refers to neuronal damage and brain physiological and structural changes caused by diabetes. Accumulating evidences indicated that diabetes is highly connected with a progressive cognitive impairment and results in AD . Hippocampus is responsible for cognitive function. Some researches have shown that type 2 diabetes results in hippocampal neuron apoptosis by Estrogen receptors-PI3/Akt pathway . Kim Ms et al.  demonstrated that microbial dysbiosis contribute to the pathogenesis of Alzheimer’s disease related to cognitive impairment and fecal microbiota transfer reversed the AD pathophysiology. Meanwhile, dysbiosis of microbiota played an important role in the pathological process of diabetic cognition impairment . The underlying mechanism needs to be explored.
Sitagliptin is currently the common treatment for type 2 diabetes through inhibiting DPP-4 activity. We observed that sitagliptin alleviated hippocampal neuron loss induced by type 2 diabetes in rats. As fore-mentioned, microbiota affects process of AD and diabetic cognition impairment. Therefore, the relationships between sitagliptin and diabetic central neuropathy, as well as microbiota remain unclear.
Type 2 diabetes is characterized by elevated glucose levels due partly to insulin resistance and insufficient secretion of insulin, with increased prevalence at an alarming rate. Genetic and environmental factors, and other risk factors containing age, dietary habits, obesity and lifestyle, have been confirmed to account for the disease. Some studies observed that dysbiosis of the human gut microbiota is connected with type 2 diabetes. Qin  found type 2 diabetes patients had gut bacterial dysbiosis with a decrease in butyrate-producing bacteria, which have been confirmed to alleviate several diseases, and a rise in some opportunistic pathogens such as Escherichia coli in Chinese population. Similar results were observed in European type 2 diabetes patients . Similarly, some researches found that gut microbiota were substantially altered in STZ-induced type 2 diabetic rats [15, 30].
The gut microbiota comprising the collective genome of 100 trillion microorganisms in the gastrointestinal tract is regarded as a major player affecting the health status of the host. Intestinal microbes influence the central nervous system through central, autonomic and enteroendocrine nervous system, production of various metabolites as well as microbial-associated-molecular patterns (MAMPs) produced by microbiota, the bidirectional communication has been suggested a microbiota-gut-brain axis . Accumulating evidences showed that disorder of gut microbiota involved in the development and function of central nervous system. Some studies showed that dysbiosis of microbiota was associated with cognitive and behavioral dysfunction in mice [31, 32]. Microbiota-gut-brain axis is regarding as a recognized targeted strategy for the prevention and treatment of central nervous system related diseases [33, 34].
Our objective was to determine whether sitagliptin treatment played an important role in returning the gut microbiota back to the normal. Moreover, the sitagliptin-treated rats displayed changed microbial diversity compared to diabetic rats, which was in accordance with the previous study . The data showed that the rat gut microbiome was largely dominated by phyla Firmicutes and Bacteroidetes. Compared to normal rats, diabetic rats related to a significant shift in the gut microbiota, with a reduced percentage of Firmicutes and Candidatus_Saccharibacteria and an increased proportion of phyla Bacteroidetes and Spirochaetes. In addition, the Bacteroidetes: Firmicutes ratio also increased in the T2DM group, similar to previous studies . Furthermore, sitagliptin treatment normalized the ratio of Bacteroidetes: Firmicutes, with an increase in abundance of Firmicutes and a decrease of Bacteroidetes, contradicting the previous findings with higher proportion of Bacteroidetes and lower proportion of Firmicutes after sitagliptin treatment . The main reason for the inconsistency was the limitation of small sample size, which should be confirmed further with larger size of samples.
Enterobacteriaceae, Lactobacillacea, Prevotellaceae, and Ruminococcaceae were the main families of all the rats. As a part of the phylum Bacteroidetes, family Prevotellaceae has some genera associated with an increase risk of diabetes mellitus . It was more abundant in the diabetic rats compared to the normal rats, decreased in the sitagliptin-treated rats in this study. Among phylum Firmicutes, family Ruminococcaceae was a major utilizer of plant polysaccharides [38, 39], and its enrichment might counteract the development of autoimmune diabetes . The result showed that a higher abundance of Ruminococcaceae in the sitagliptin-treated rats than in the diabetic rats. Notably, family Lactobacillaceae increased significantly after sitagliptin treatment compared to the T2DM group, one of its main genera, Lactobacillus also presented a higher abundance in the T2DM-Sit group according to the LEfSe analysis. The mechanism might be related that some Lactobacillus species could metabolize tryptophan into indole-3-aldyhyde, which acts on the aryl hydrocarbon receptor (AhR) in intestinal immune cells . The ligand for AhR was indole, which could trigger the secretion of glucagon-like peptide -1 in intestinal L cells .
The LEfSe analysis showed that the diabetic rats with an enrichment of family Enterobacteriaceae and Enterococcaceae, order Enterobacteriales. As potential pro-inflammatory microorganism in the gut, Enterobacteriaceae and Enterococcaceae both presented an increase in the T2DM patients . They could further contribute to the rise of imflammatory level in the host, supporting the evolution of insulin resistance . Previous study also observed a higher abundance of the gram-negative bacteria Enterobacteriales in the diabetic condition, which may be mechanistically linked to increased colonic permeability irrespective of glucose tolerance status .
In our study, genus Lactobacillus, genera Streptococcus, Clostridium and Fusobacterium increased significantly compared to diabetic group after sitagliptin treatment. They can produce a range of neurotransmitters to participate in the regulation of varied and important physiological processes, including immunomodulation, adiposity, and energy balance , genus Streptococcus produce serotonin . Lactobacillus produces acetylcholine and γ-aminobutyric acid . Species Clostridium sporogens of genus Clostridium metabolizes tryptophan into indole and subsequently 3-indolepropionic acid, a highly potent neuroprotective antioxidant . Moreover, Clostridium and Fusobacterium are SCFAs-producing bacterium mainly exists in cecum and colon .
Short-chain fatty acids (SCFAs) including acetic acid, butyric acid and propionic acid are major metabolic and products of gut microbial degradation of dietary fiber in the colon . SCFAs are ligands for G protein-coupled receptor (GPCR) GPR41 and induce secretion of the enteroendocrine hormone peptide YY (PYY) from gut epithelial L-cells, which restrains gut motility and add energy harvest from the diet . Accumulating evidence showed that SCFA-GPR interactions would allow direct signaling from gut to the central nervous system. The maturation and function of microglia, which are the resident macrophages of the brain, were dependent on gut microbiota, and SCFAs and GPR were necessary to maintain microglia homeostasis and integrity of the blood-brain barrier in mice [51, 52]. Recently, a study showed that high-fructose diet-induced gut dysbiosis with reduced SCFA in the impairment of colonic epithelia barrier that resulted in hippocampal neuroinflammation and neuronal loss in mice, and the neurodegenerative changes could be prevented by SCFA treatment . These results further suggest that SCFAs exert protective effects on modulation of neurological function. Which may help to explain partly the neuroprotective effects of sitagliptin on hippocamlal neuron loss induced by type 2 diabetes by triggering microbiota-gut-brain axis.
However, lack of the proportions of SCFAs and neurotransmitters existed in this study, which made the changes of SCFAs after sitagliptin treatment without data support. Therefore in the further study, the quantification of these metabolites should be taken into account. Additionally, there are some issues that still need to be resolved: detection of microbial interaction and discrepancy at lower taxonomic levels; large cohort studies to confirm these findings; longitudinal studies to track the gut microbiome and its influences on CNS function.