Background Cachexia is a life-threatening condition observed in several pathologies, such as cancer or chronic diseases. Interleukin 10 (Il10) gene transfer is known to improve cachexia by downregulating Il6. Here, we used an IL10-knockout mouse model to simulate cachexia and investigate the effects of eggshell membrane (ESM), a resistant protein, on general pre-cachexia symptoms, which is particularly important for the development of cachexia therapeutics. Methods Five-week-old male C57BL6/J mice were fed an AIN-93G powdered diet (WT), and 5-week-old male B6.129P2-Il10 < tm1Cgn>/J (IL10 À/À ) mice were fed either the AIN-93G diet (KO) or an 8% ESM-containing diet (KOE) for 28 weeks. The tissue weight and levels of anaemia-, blood glucose-, lipid metabolism-, and muscular and colonic inflammation-related biochemical markers were measured. Transcriptomic analysis on liver and colon mucus and proteomic analysis on skeletal muscle were performed. Ingenuity Pathway Analysis was used to identify molecular pathways and networks. Caecal short-chain fatty acids (SCFAs) were identified using HPLC, and caecal bacteria DNA were subjected to metagenomic analysis. Flow cytometry analysis was performed to measure the CD4 + IL17 + T cells in mesenteric lymph nodes. ResultsThe body weight, weight of gastrocnemius muscle and fat tissues, colon weight/length ratio, plasma HDL and NEFA, muscular PECAM-1 levels (P < 0.01), plasma glucose and colonic mucosal myeloperoxidase activity (P < 0.05) and T helper (Th) 17 cell abundance (P = 0.071) were improved in KOE mice over KO mice. Proteomic analysis indicated the protective role of ESM in muscle weakness and maintenance of muscle formation (>1.5-fold). Transcriptomic analysis revealed that ESM supplementation suppressed the LPS/IL1-mediated inhibition of RXR function pathway in the liver and downregulated the colonic mucosal expression of chemokines and Th cell differentiation-related markers (P < 0.01) by suppressing the upstream BATF pathway. Analysis of the intestinal microenvironment revealed that ESM supplementation ameliorated the microbial alpha diversity and the abundance of microbiota associated with the degree of inflammation (P < 0.05) and increased the level of total organic acids, particularly of SCFAs such as butyrate (2.3fold), which could inhibit Th1 and Th17 production. Conclusions ESM supplementation ameliorated the chief symptoms of cachexia, including anorexia, lean fat tissue mass, skeletal muscle wasting and reduced physical function. ESM also improved colon and skeletal muscle inflammation, lipid metabolism and microbial dysbiosis. These results along with the suppressed differentiation of Th cells could be associated with the beneficial intestinal microenvironment and, subsequently, attenuation of pre-cachexia. Our findings provide insights into the potential of ESM in complementary interventions for pre-cachexia prevention.
Background: Aquaporin 5 (AQP5) is a water channel-forming protein that plays a key role in saliva secretion. A decrease in masticatory function associated with the molar extraction adversely affects the submandibular salivary gland (SMG) in rats, inducing hypertrophic changes in the acinar cells and the expression of AQP5 in acinar cells or intercalated duct of the SMG. However, changes in AQP5 expression and localization in the SMG in association with occlusal modification have not been fully characterized. Methods: We examined the influence of the decline and recovery of masticatory function on expression and localization of AQP5 in the rat SMG by inserting and removing an incisor bite plate (IBP). Thirty 5-week-old male Wistar rats were randomly divided into IBP (n = 12), recovery (REC) (n = 6), and control (CON) (n = 12) groups. Each rat in both the IBP and REC groups was fitted with the IBP on its maxillary incisors. Rats without the IBPs served as controls. All rats were fed powder diet and water ad libitum. Rats in the IBP and CON groups were sacrificed after 14 (n = 6) and 28 (n = 6) days after the IBP attachment. In the REC group, the IBP was detached on the 14th day and sacrificed on 28th day after the IBP attachment. AQP5 mRNA expression was quantified by reverse transcription-polymerase chain reaction. Changes in the localization of AQP5 were tracked by immunohistochemical staining. Results: Attachment of IBP resulted in a decrease in the expression of AQP5 in the IBP group. Changes in the localization of AQP5 were observed between 14 and 28 days in the IBP group. In contrast, changes in the expression and localization of AQP5 were not observed in the REC group. Conclusion: Findings suggested that a loss of molar occlusion, due to the IBP attachment, altered AQP5 expression and localization in the rat SMG. However, removal of the bite plate allowed the recovery of both AQP5 expression and its normal localization in the SMGs.
Objectives: Ghrelin is a key regulator of food intake and is considered a hunger hormone that affects cognition, memory, glucose metabolism, and antidepressant effects. Altered occlusion, such as a loss of molars, has been thought to retard digestive function. However, the association between occlusion and digestive function remains poorly understood. Here, we aimed to explore the effect of bilateral maxillary molar extraction on the gastrointestinal mucosa of growing rats and the expression of ghrelin and its receptor, growth hormone secretagogue receptor (GHSR). Material and Methods: Twenty-four male 5-week-old Wistar rats were divided into control (CON) and experimental (EXP) groups (n = 12/group). The rats in the EXP group underwent extraction of the bilateral maxillary first, second, and third molars under general anesthesia. Rats in the CON group underwent a sham operation. All rats in both the CON and EXP groups were fed a powder diet and water ad libitum. The body weight of all rats was monitored throughout the EXP period. Rats in both the CON and EXP groups were euthanized on days 14 and 28, and the stomachs were isolated and subjected to histological analysis. Paraffin serial sections were prepared using a microtome for hematoxylin-eosin and immunohistochemical staining using anti-ghrelin and anti-GHSR antibodies. The distribution and expression of ghrelin-immunopositive and GHSR cells were detected and observed under a light microscope. Data were statistically analyzed using t-tests (P < 0.05). Results: There were no significant differences in body weight between the CON and EXP groups throughout the EXP period. Histological analysis showed that the area of the submucosa (ASM), and the number of ghrelinimmunopositive cells were significantly decreased in the EXP group compared with the CON group on day 14. Alternatively, there was no significant difference in the ASM and the number of ghrelin-immunopositive cells between the CON and EXP groups on day 28, whereas the number of ghrelin receptors showed no differences across groups. Furthermore, the number of eosinophilic blood cells significantly increased in the EXP group on days 14 and 28. Conclusion: Our findings suggest that bilateral maxillary molar extraction may trigger stomach mucosal changes and alter digestive function through ghrelin expression in rats. This is the first report that occlusal deficiency could alter ghrelin expression in the mucosa of the rat stomach, thus raising concerns about the consequential role of ghrelin.
OBJECTIVES:To determine whether the modification of dental occlusion, without molar extraction, affected the gustatory papillae located in the tongue of growing rats.MATERIALS AND METHODS:Five-week-old male Wistar rats were randomly divided into an anterior bite plate (ABP) group and a control group. Under general anesthesia, ABPs were placed on the occlusal surfaces of the maxillary incisors, while metal caps covered the mandibular incisal edges of the rats in the ABP group. The control group rats underwent a sham operation. The rats in both groups were euthanized 14 days after the procedure. The circumvallate papillae and taste buds were analyzed by immunohistochemical methods, and the fungiform papillae were observed and counted after immersion of the tongue in 1% methylene blue.RESULTS:Two weeks after ABP insertion and mandibular incisal cap placement, the gustatory papillae exhibited morphological and structural changes. The rats in the ABP group had exhibited significantly fewer fungiform papillae, and narrower circumvallate papillae, with greater trench depths, larger trench profile areas, smaller taste bud profile areas, lower ratios of the taste bud profile area to the trench profile area, and more taste buds than those in the control group.CONCLUSIONS:Our findings support the association between occlusal and taste functions and provide a basis for further studies on the gustatory function. In conclusion, loss of molar occlusion, resulting from the ABP and metal cap insertion, altered the peripheral gustatory receptors in the growing rats.
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