This study aims to investigate the mechanism of Qingrun Decoction in alleviating hepatic insulin resistance in type 2 diabetes mellitus(T2DM) rats through the reprogramming of amino acid metabolism. A T2DM rat model was established by inducing insulin resistance through a high-fat diet combined with intraperitoneal injection of streptozotocin. The model rats were randomly divided into five groups: model group, high-, medium-, and low-dose Qingrun Decoction groups, and metformin group. A normal control group was also established. The rats in the normal and model groups received 10 mL·kg~(-1) distilled water daily by gavage. The metformin group received 150 mg·kg~(-1) metformin suspension by gavage, and the Qingrun Decoction groups received 11.2, 5.6, and 2.8 g·kg~(-1) Qingrun Decoction by gavage for 8 weeks. Blood lipid levels were measured in different groups of rats. Pathological damage in rat liver tissue was assessed by hematoxylin-eosin(HE) staining and oil red O staining. Transcriptome sequencing and untargeted metabolomics were performed on rat liver and serum samples, integrated with bioinformatics analyses. Key metabolites(branched-chain amino acids, BCAAs), amino acid transporters, amino acid metabolites, critical enzymes for amino acid metabolism, resistin, adiponectin(ADPN), and mammalian target of rapamycin(mTOR) pathway-related molecules were quantified using quantitative real-time polymerase chain reaction(qRT-PCR), Western blot, and enzyme-linked immunosorbent assay(ELISA). The results showed that compared with the normal group, the model group had significantly increased serum levels of total cholesterol(TC), triglycerides(TG), low-density lipoprotein cholesterol(LDL-C), and resistin and significantly decreased ADPN levels. Hepatocytes in the model group exhibited loose arrangement, significant lipid accumulation, fatty degeneration, and pronounced inflammatory cell infiltration. In liver tissue, the mRNA transcriptional levels of solute carrier family 7 member 2(Slc7a2), solute carrier family 38 member 2(Slc38a2), solute carrier family 38 member 4(Slc38a4), and arginase(ARG) were significantly downregulated, while the mRNA transcriptional levels of solute carrier family 1 member 4(Slc1a4), solute carrier family 16 member 1(Slc16a1), and methionine adenosyltransferase(MAT) were upregulated. Furthermore, the mRNA transcription and protein expression levels of branched-chain α-keto acid dehydrogenase E1α(BCKDHA) and DEP domain-containing mTOR-interacting protein(DEPTOR) were downregulated, while mRNA transcription and protein expression levels of mTOR, as well as ribosomal protein S6 kinase 1(S6K1), were upregulated. The levels of BCAAs and S-adenosyl-L-methionine(SAM) were elevated. The serum level of 6-hydroxymelatonin was significantly reduced, while imidazole-4-one-5-propionic acid and N-(5-phospho-D-ribosyl)anthranilic acid levels were significantly increased. Compared with the model group, Qingrun Decoction significantly reduced blood lipid and resistin levels while increasing ADPN levels. Hepatocytes had improved morphology with reduced inflammatory cells, and fatty degeneration and lipid deposition were alleviated. Differentially expressed genes and differential metabolites were mainly enriched in amino acid metabolic pathways. The expression levels of Slc7a2, Slc38a2, Slc38a4, and ARG in the liver tissue were significantly upregulated, while Slc1a4, Slc16a1, and MAT expression levels were significantly downregulated. BCKDHA and DEPTOR expression levels were upregulated, while mTOR and S6K1 expression levels were downregulated. Additionally, the levels of BCAAs and SAM were significantly decreased. The serum level of 6-hydroxymelatonin was increased, while those of imidazole-4-one-5-propionic acid and N-(5-phospho-D-ribosyl)anthranilic acid were decreased. In summary, Qingrun Decoction may improve amino acid metabolism reprogramming, inhibit mTOR pathway activation, alleviate insulin resistance in the liver, and mitigate pathological damage of liver tissue in T2DM rats by downregulating hepatic BCAAs and SAM and regulating key enzymes involved in amino acid metabolism, such as BCKDHA, ARG, and MAT, as well as amino acid metabolites and transporters.
Animals ; Drugs, Chinese Herbal/administration & dosage* ; Rats ; Insulin Resistance ; Diabetes Mellitus, Type 2/genetics* ; Male ; Liver/drug effects* ; Amino Acids/metabolism* ; Rats, Sprague-Dawley ; Humans ; Metabolic Reprogramming

OBJECTIVE: To identify and characterize read-through RNAs and read-through circular RNAs (rt-circ-HS) derived from transcriptional read-through hypoxia inducible factor 1α (HIF1α) and small nuclear RNA activating complex polypeptide 1 (SNAPC1) the two adjacent genes located on chromosome 14q23, in renal carcinoma cells and renal carcinoma tissues, and to study the effects of rt-circ-HS on biological behavior of renal carcinoma cells and on regulation of HIF1α. METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) and Sanger sequencing were used to examine expression of read-through RNAs HIF1α-SNAPC1 and rt-circ-HS in different tumor cells. Tissue microarrays of 437 different types of renal cell carcinoma (RCC) were constructed, and chromogenic in situ hybridization (ISH) was used to investigate expression of rt-circ-HS in different RCC types. Small interference RNA (siRNA) and artificial overexpression plasmids were designed to examine the effects of rt-circ-HS on 786-O and A498 renal carcinoma cell proliferation, migration and invasiveness by cell counting kit 8 (CCK8), EdU incorporation and Transwell cell migration and invasion assays. RT-PCR and Western blot were used to exa-mine expression of HIF1α and SNAPC1 RNA and proteins after interference of rt-circ-HS with siRNA, respectively. The binding of rt-circ-HS with microRNA 539 (miR-539), and miR-539 with HIF1α 3' untranslated region (3' UTR), and the effects of these interactions were investigated by dual luciferase reporter gene assays. RESULTS: We discovered a novel 1 144 nt rt-circ-HS, which was derived from read-through RNA HIF1α-SNAPC1 and consisted of HIF1α exon 2-6 and SNAPC1 exon 2-4. Expression of rt-circ-HS was significantly upregulated in 786-O renal carcinoma cells. ISH showed that the overall positive expression rate of rt-circ-HS in RCC tissue samples was 67.5% (295/437), and the expression was different in different types of RCCs. Mechanistically, rt-circ-HS promoted renal carcinoma cell proliferation, migration and invasiveness by functioning as a competitive endogenous inhibitor of miR-539, which we found to be a potent post-transcriptional suppressor of HIF1α, thus promoting expression of HIF1α. CONCLUSION The novel rt-circ-HS is highly expressed in different types of RCCs and acts as a competitive endogenous inhibitor of miR-539 to promote expression of its parental gene HIF1α and thus the proliferation, migration and invasion of renal cancer cells.
Humans ; Carcinoma, Renal Cell/pathology* ; Cell Proliferation ; Hypoxia ; Kidney Neoplasms ; MicroRNAs/genetics* ; Neoplasm Invasiveness/genetics* ; RNA, Circular/metabolism* ; RNA, Small Interfering ; Hypoxia-Inducible Factor 1, alpha Subunit/genetics*

The mammalian epididymis not only plays a fundamental role in the maturation of spermatozoa, but also provides protection against various stressors. The foremost among these is the threat posed by oxidative stress, which arises from an imbalance in reactive oxygen species and can elicit damage to cellular lipids, proteins, and nucleic acids. In mice, the risk of oxidative damage to spermatozoa is mitigated through the expression and secretion of glutathione peroxidase 5 (GPX5) as a major luminal scavenger in the proximal caput epididymidal segment. Accordingly, the loss of GPX5-mediated protection leads to impaired DNA integrity in the spermatozoa of aged Gpx5

Activation of acid-sensing ion channels (ASICs) plays an important role in neuroinflammation.Macrophage recruitment to the sites of inflammation is an essential step in host defense.ASIC 1 and ASIC3 have been reported to mediate the endocytosis and maturation of bone marrow derived macrophages.However,the expression and inflammation-related functions of ASICs in RAW 264.7 cells,another common macrophage,are still elusive.In the present study,we first demonstrated the presence of ASIC 1,ASIC2a and ASIC3 in RAW 264.7 macrophage cell line by using reverse transcriptase polymerase chain reaction (RT-PCR),Western blotting and immunofluorescence experiments.The non-specific ASICs inhibitor amiloride and specific homomeric ASIC 1 a blocker PcTx 1 reduced the production of iNOS and COX-2 by LPS-induced activating RAW 264.7 cells.Furthermore,not only amiloride but also PcTx 1 inhibited the migration and LPS-induced apoptosis of RAW 264.7 cells.Taken together,our findings suggest that ASICs promote the inflammatory response and apoptosis of RAW 264.7 cells,and ASICs may serve as a potential novel target for immunological disease therapy.

Objective To investigate the role of BMSCg on enhancing the implant survival and bacheal epithelium regeneration. Methods After transplanted with cryopreserved 2 weeks and 6 weeks allocraft, PHK-26 labeled 3-5 passage BMSCs were injected into the recipient rats via tail vein. Rats in the control groups were injected with the same amount of PBS.We observed the histology of the transplanted trachea including epithelium growth and regeneration, and the PKH-26 fluorescence levels at the para-anastomotic trachea to evaluate the role of BMSC transplantation on the epithelium regeneration. Results Rats from BMSCs injection group survived a long period. Histological observation showed that the tracheal lumen was covered by psudo-striated ciliated columnar epithelium. The cartilage structure was intact. There are no signs of denaturation and necrosis. In the PBS injection group, epithelium regeneration is better in PBS-6-week group than PBS-2-week group. The longest survival time in PBS-6-week group was 32 days, whereas it was 10 days in PBS-2-week group. In BMSCs injection group, rats in BMSC-6-week groups survived longer than 8-week group(12 rats were terminated at 1 week, 4 weeks and 8weeks as planned). There was one rat who survived and were terminated at the designated 8 weeks time point (there were 8regenerated epithelium was similar in the two BMSC transplanted groups. PKH-26 labeled BMSCs migrated to the implant site and showed red fluorescence, with most red fluorescence shown at the anastomotic part. Conclusion BMSCs can migrate to the impaired tissue to repair it. BMSCs may exert their reparation function via enhancing epithelium regeneration.

OBJECTIVETo assess the effect of angiotensin II type 1 (AT(1)) receptor antagonist losartan on myocardium connexin43 (Cx43) gap junction (GJ) expression in spontaneously hypertensive rats (SHRs) and investigate possible mechanisms.

METHODSSixteen 9-week-old male SHRs and 8 age-matched male Wistar-Kyoto (WKY) rats were included in this study. SHRs were randomly divided into two groups to receive losartan at 30 mg/(kg x d) by oral gavage once daily for 8 weeks (SHR-L) or vehicle (0.9% saline) to act as controls (SHR-V); WKY rats receiving vehicle for 8 weeks served as normotensive controls. At the end of the experiment, rats were sacrificed and the hearts were removed. Expressions of Cx43 and nuclear factor-kappaB p65 (NF-kappaB p65) proteins in all three groups were observed and further investigations on the effect of angiotensin II type 1 receptor antagonist losartan (30 mg/(kg x d), 8 weeks) on Cx43 expression were conducted with Western blot and immunohistochemistry. NF-kappaB p65 protein in nuclear extracts was determined by Western blot.

RESULTSLeft ventricular (LV) hypertrophy was prominent in SHRs, Cx43 and NF-kappaB p65 protein expressions were obviously upregulated and Cx43 distribution was dispersed over the cell surface. Treatment with losarton reduced the over-expressions of Cx43 and NF-kappaB p65 in LV myocardium. The distribution of Cx43 gap junction also became much regular and confined to intercalated disk after losartan treatment.

CONCLUSIONCx43 level was upregulated in LV myocardium of SHR during early stage of hypertrophy. Angiotensin II type 1 receptor antagonist losartan prevented Cx43 gap junction remodeling in hypertrophied left ventricles, possibly through the NF-kappaB pathway.

Angiotensin II Type 1 Receptor Blockers ; pharmacology ; Animals ; Blood Pressure ; drug effects ; Blotting, Western ; Connexin 43 ; metabolism ; Hypertension ; drug therapy ; metabolism ; physiopathology ; Hypertrophy, Left Ventricular ; drug therapy ; metabolism ; pathology ; Losartan ; pharmacology ; Male ; Myocardium ; metabolism ; Natriuretic Peptide, Brain ; blood ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY ; Transcription Factor RelA ; metabolism

Objective To understand the epidemiological characteristics of age distribution of measles and related policies on measles vaccines (live; MV) in infants through analyzing the antibody levels of comparison in maternal-infant pairs. Transition of immunity in infants was also studied to provide theoretic basis for measles immunization strategy and to reduce the incidence of month-old infants. Methods In cities of Ningbo, Harbin, and Jinan from Zhejiang, Heilongiiang and Shandong provinces, data was collected from 2004 to 2007 and analyzed regarding the epidemic situation of measles. Studies on maternal-transferred measles antibody were carried our sero-epidemiologically. Results Most of the measles cases were found among babies younger than 12 months,and the incidence of < 1 year olds had been increasing.The distribution was dominated by 5-8 month olds in infant measles cases. The positive rate and GRMT of measles antibody in newborns were 89.3 percent and 738.93. The positive rate of the measles antibody and GMRT of the 6-month infant were 6.9 % and 6.89, while 6.7 % and 3.69 in 8-month infant. There wasa declining trend of the positive rate of the measles antibody during the newborns to 8-month infant. The positive rate and GRMT of measles antibody in mothers were 84.3 percent and 516.94. Mother's measles antibodies mainly to be at low and moderate level, which accounted for 50.4 percent and 30.3 percent respectively, the correlation coefficient between mother and infant was 0. 840. Conclusion Maternal-transferred measles antibody decreased as the growth of infants. The positive rates of measles antibody were quite low in 6-month and 8-month olds which were the age range that needs most attention.