Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Background: In acute and chronic renal inflammatory diseases, the activation of inflammatory cells is involved in the defect of erythropoietin (EPO) production. Ras guanine nucleotide-releasing protein-4 (RasGRP4) promotes renal inflammatory injury in type 2 diabetes mellitus (T2DM). Our study aimed to investigate the role and mechanism of RasGRP4 in the production of renal EPO in diabetes. Methods: The degree of tissue injury was observed by pathological staining. Inflammatory cell infiltration was analyzed by immunohistochemical staining. Serum EPO levels were detected by enzyme-linked immunosorbent assay, and EPO production and renal interstitial fibrosis were analyzed by immunofluorescence. Quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of key inflammatory factors and the activation of signaling pathways. In vitro, the interaction between peripheral blood mononuclear cells (PBMCs) and C3H10T1/2 cells was investigated via cell coculture experiments. Results: RasGRP4 decreased the expression of hypoxia-inducible factor 2-alpha (HIF2A) via the ubiquitination–proteasome degradation pathway and promoted myofibroblastic transformation by activating critical inflammatory pathways, consequently reducing the production of EPO in T2DM mice. Conclusion RasGRP4 participates in the production of renal EPO in diabetic mice by affecting the secretion of proinflammatory cytokines in PBMCs, degrading HIF2A, and promoting the myofibroblastic transformation of C3H10T1/2 cells.

Objective To analyze the genetic subtypes of human immunodeficiency virus (HIV) and the prevalence of pretreatment drug resistance in the newly reported HIV-infected men in Guangxi. Methods The stratified random sampling method was employed to select the newly reported HIV-infected men aged≥50 years old in 14 cities of Guangxi from January to June in 2020.The pol gene of HIV-1 was amplified by nested reverse transcription polymerase chain reaction and then sequenced.The mutation sites associated with drug resistance and the degree of drug resistance were then analyzed. Results A total of 615 HIV-infected men were included in the study.The genetic subtypes of CRF01_AE,CRF07_BC,and CRF08_BC accounted for 57.4% (353/615),17.1% (105/615),and 22.4% (138/615),respectively.The mutations associated with the resistance to nucleoside reverse transcriptase inhibitors (NRTI),non-nucleoside reverse transcriptase inhibitors (NNRTI),and protease inhibitors occurred in 8 (1.3%),18 (2.9%),and 0 patients,respectively.M184V (0.7%) and K103N (1.8%) were the mutations with the highest occurrence rates for the resistance to NRTIs and NNRTIs,respectively.Twenty-two (3.6%) patients were resistant to at least one type of inhibitors.Specifically,4 (0.7%),14 (2.3%),4 (0.7%),and 0 patients were resistant to NRTIs,NNRTIs,both NRTIs and NNRTIs,and protease inhibitors,respectively.The pretreatment resistance to NNRTIs had much higher frequency than that to NRTIs (2.9% vs.1.3%;χ²=3.929,P=0.047).The prevalence of pretreatment resistance to lamivudine,zidovudine,tenofovir,abacavir,rilpivirine,efavirenz,nevirapine,and lopinavir/ritonavir was 0.8%, 0.3%, 0.7%, 1.0%, 1.3%, 2.8%, 2.9%, and 0, respectively. Conclusions CRF01_AE,CRF07_BC,and CRF08_BC are the three major strains of HIV-infected men≥50 years old newly reported in Guangxi,2020,and the pretreatment drug resistance demonstrates low prevalence.
Male ; Humans ; Middle Aged ; Reverse Transcriptase Inhibitors/therapeutic use* ; HIV Infections/drug therapy* ; Drug Resistance, Viral/genetics* ; China/epidemiology* ; Mutation ; HIV-1/genetics* ; Protease Inhibitors/therapeutic use* ; Genotype

Monocytes are key effectors in autoimmunity-related diseases in the central nervous system (CNS) due to the critical roles of these cells in the production of proinflammatory cytokines, differentiation of T-helper (Th) cells, and antigen presentation. The JAK-STAT signaling is crucial for initiating monocytes induced immune responses by relaying cytokines signaling. However, the role of this pathway in modulating the communication between monocytes and Th cells in the pathogenesis of multiple sclerosis (MS) is unclear. Here, we show that the JAK1/2/3 and STAT1/3/5/6 subtypes involved in the demyelination mediated by the differentiation of pathological Th1 and Th17 and the CNS-infiltrating inflammatory monocytes in experimental autoimmune encephalomyelitis (EAE), a model for MS. JAK inhibition prevented the CNS-infiltrating CCR2-dependent Ly6C^hi monocytes and monocyte-derived dendritic cells in EAE mice. In parallel, the proportion of GM-CSF⁺CD4⁺ T cells and GM-CSF secretion were decreased in pathological Th17 cells by JAK inhibition, which in turns converted CNS-invading monocytes into antigen-presenting cells to mediate tissue damage. Together, our data highlight the therapeutic potential of JAK inhibition in treating EAE by blocking the GM-CSF-driven inflammatory signature of monocytes.

Objective: To analyze the clinicopathological features and gene mutations of primary gastrointestinal stromal tumors (GISTs) of the stomach and intestine and the prognosis of intermediate- and high-risk GISTs. Methods: This was a retrospective cohort study. Data of patients with GISTs admitted to Tianjin Medical University Cancer Institute and Hospital from January 2011 to December 2019 were collected retrospectively. Patients with primary gastric or intestinal disease who had undergone endoscopic or surgical resection of the primary lesion and were confirmed pathologically as GIST were included. Patients treated with targeted therapy preoperatively were excluded. The above criteria were met by 1061 patients with primary GISTs, 794 of whom had gastric GISTs and 267 intestinal GISTs. Genetic testing had been performed in 360 of these patients since implementation of Sanger sequencing in our hospital in October 2014. Gene mutations in KIT exons 9, 11, 13, and 17 and PDGFRA exons 12 and 18 were detected by Sanger sequencing. The factors investigated in this study included: (1) clinicopathological data, such as sex, age, primary tumor location, maximum tumor diameter, histological type, mitotic index (/5 mm²), and risk classification; (2) gene mutation; (3) follow-up, survival, and postoperative treatment; and (4) prognostic factors of progression-free survival (PFS) and overall survival (OS) for intermediate- and high-risk GIST. Results: (1) Clinicopathological features: The median ages of patients with primary gastric and intestinal GIST were 61 (8-85) years and 60 (26-80) years, respectively; The median maximum tumor diameters were 4.0 (0.3-32.0) cm and 6.0 (0.3-35.0) cm, respectively; The median mitotic indexes were 3 (0-113)/5 mm² and 3 (0-50)/5 mm², respectively; The median Ki-67 proliferation indexes were 5% (1%-80%) and 5% (1%-50%), respectively. The rates of positivity for CD117, DOG-1, and CD34 were 99.7% (792/794), 99.9% (731/732), 95.6% (753/788), and 100.0% (267/267), 100.0% (238/238), 61.5% (163/265), respectively. There were higher proportions of male patients (χ²=6.390, P=0.011), tumors of maximum diameter > 5.0 cm (χ²=33.593, P<0.001), high-risk (χ²=94.957, P<0.001), and CD34-negativity (χ²=203.138, P<0.001) among patients with intestinal GISTs than among those with gastric GISTs. (2) Gene mutations: Gene mutations were investigated in 286/360 patients (79.4%) with primary gastric GISTs and 74/360 (20.6%) with primary intestinal GISTs. Among the 286 patients with gastric primary GISTs, 79.4% (227/286), 8.4% (24/286), and 12.2% (35/286), had KIT mutations, PDGFRA mutations, and wild-type, respectively. Among the 74 patients with primary intestinal GISTs, 85.1% (63/74) had KIT mutations and 14.9% (11/74) were wild-type. The PDGFRA mutation rate was lower in patients with intestinal GISTs than in those with gastric GISTs[ 0% vs. 8.4%(24/286), χ²=6.770, P=0.034], whereas KIT exon 9 mutations occurred more often in those with intestinal GISTs [22.2% (14/63) vs. 1.8% (4/227), P<0.001]. There were no significant differences between gastric and intestinal GISTs in the rates of KIT exon 11 mutation type and KIT exon 11 deletion mutation type (both P>0.05). (3) Follow-up, survival, and postoperative treatment: After excluding 228 patients with synchronous and metachronous other malignant tumors, the remaining 833 patients were followed up for 6-124 (median 53) months with a follow-up rate of 88.6% (738/833). None of the patients with very low or low-risk gastric (n=239) or intestinal GISTs (n=56) had received targeted therapy postoperatively. Among 179 patients with moderate-risk GISTs, postoperative targeted therapy had been administered to 88/155 with gastric and 11/24 with intestinal GISTs. Among 264 patients with high-risk GISTs, postoperative targeted therapy had been administered to 106/153 with gastric and 62/111 with intestinal GISTs. The 3-, 5-, and 10-year PFS of patients with gastric or intestinal GISTs were 96.5%, 93.8%, and 87.6% and 85.7%, 80.1% and 63.3%, respectively (P<0.001). The 3-, 5-, and 10-year OS were 99.2%, 98.8%, 97.5% and 94.8%, 92.1%, 85.0%, respectively (P<0.001). (4) Analysis of predictors of intermediate- and high-risk GISTs: The 5-year PFS of patients with gastric and intestinal GISTs were 89.5% and 73.2%, respectively (P<0.001); The 5-year OS were 97.9% and 89.3%, respectively (P<0.001). Multivariate analysis showed that high risk (HR=2.918, 95%CI: 1.076-7.911, P=0.035) and Ki-67 proliferation index > 5% (HR=2.778, 95%CI: 1.389-5.558, P=0.004) were independent risk factors for PFS in patients with intermediate- and high-risk GISTs (both P<0.05). Intestinal GISTs (HR=3.485, 95%CI: 1.407-8.634, P=0.007) and high risk (HR=3.753,95%CI:1.079-13.056, P=0.038) were independent risk factors for OS in patients with intermediate- and high-risk GISTs (both P<0.05). Postoperative targeted therapy was independent protective factor for PFS and OS (HR=0.103, 95%CI: 0.049-0.213, P<0.001; HR=0.210, 95%CI:0.078-0.564,P=0.002). Conclusions: Primary intestinal GIST behaves more aggressively than gastric GISTs and more frequently progress after surgery. Moreover, CD34 negativity and KIT exon 9 mutations occur more frequently in patients with intestinal GISTs than in those with gastric GISTs.
Male ; Humans ; Gastrointestinal Stromal Tumors/surgery* ; Retrospective Studies ; Ki-67 Antigen ; Stomach Neoplasms/pathology* ; Prognosis ; Mutation ; Intestines/pathology* ; Proto-Oncogene Proteins c-kit/genetics* ; Receptor, Platelet-Derived Growth Factor alpha/genetics*

OBJECTIVE: To observe the effect of transcutaneous auricular vagus nerve stimulation (taVNS) on the sleep quality and nocturnal heart rate variability (HRV) in patients with primary insomnia. METHODS: Twenty-one patients with primary insomnia were included. Using SDZ-ⅡB electric acupuncture apparatus, Xin (CO₁₅) and Shen (CO₁₀) were stimulated with disperse-dense wave, 4 Hz/ 20 Hz in frequency, (0.2±30%) ms of pulse width and tolerable intensity. Electric stimulation was given once every morning and evening of a day, 30 min each time, for 4 weeks totally. Before and after treatment, the score of Pittsburgh sleep quality index (PSQI), objective sleep structure (total sleep time [TST], sleep latency [SL], wake after sleep onset [WASO], sleep efficiency [SE], the percentages of non-rapid eye movement period 1, 2, 3, and the percentage of rapid eye movement period to TST [N1%, N2%, N3%, REM%] ) and nocturnal HRV (high frequency [HF], low frequency [LF], the ratio of LF to HF [LF/HF], standard deviation for the normal RR intervals [SDNN], squared root of the mean sum of squares of differences between adjacent intervals RR [RMSSD], the percentage of adjacent RR intervals with differences larger than 50 ms in the entire recording [PNN50%], the mean of sinus RR intervals [NNMean] ) were compared in the patients separately. RESULTS: After treatment, the score of each item and the total score of PSQI and SL were all reduced as compared with those before treatment (P<0.01, P<0.001); SE, N3%, LF, HF, LF/HF, SDNN, NNMean and RMSSD were all increased compared with those before treatment (P<0.001, P<0.01). CONCLUSION The taVNS improves the sleep quality and objective sleep structure in patients with primary insomnia, which is probably related to the regulation of autonomic nervous functions.
Heart Rate/physiology* ; Humans ; Sleep/physiology* ; Sleep Initiation and Maintenance Disorders/therapy* ; Vagus Nerve ; Vagus Nerve Stimulation

OBJECTIVE：To screen the effective compo nent in antioxi dant active fraction of Pueraria lobata . METHODS ：The antioxidant active fraction sample （S1-S20） of 20 batches of P. lobata were prepared. HPLC method was adopted. The determination was performed on SepaxBio-C 18 column with mobile phase consisted of methanol-water （gradient elution ）at the flow rate of 0.6 mL/min. The column temperature was set at 25 ℃，and detection wavelength was set at 250 nm. HPLC fingerprints of 20 batches of P. lobata were established by the Similarity Evaluation System of TCM Chromatographic Fingerprints （2012 edition），and common peaks were identified. Cluster analysis ，principal component analysis （PCA）and orthogonal partial least squares discriminant analysis （OPLS-DA）were used to screen the effective components in antioxidant active fraction of P. lobata . RESULTS：There were 18 common peaks in HPLC fingerprints of 20 batches of antioxidant active fraction in P. lobata ，and the similarity was more than 0.99. Eight common peaks were identified ，which were 3′-hydroxypuerarin（peak 2），puerarin（peak 3）， 3′-methoxypuerarin（peak 4），daidzein（peak 5），genistein（peak 7），formononetin（peak 11），daidzein（peak 13）and genistein （peak 16）. The results of cluster analysis and PCA analysis showed that samples S 1，S3，S4，S6，S8，S18 and S 19 were clustered into one category ，and samples S 2，S5，S7，S9-S17 and S 20 were clustered into one category ；peak 2，peak 3，peak 10，peak 11 and peak 13 had great influence on principal component 1；peak 8 and peak 9 had great influence on principal component 2. OPLS-DA analysis showed that peak 4，peak 3，peak 2，peak 16，peak 13 and peak 11 had great influence on the quality of antioxidant active fraction of P. lobata . CONCLUSIONS ： HPLC fingerprint for active fraction of P. lobata is established in the study and 8 components are identified ；among them ， com puerarin，3′-hydroxypuerarin，daidzein and formononetin maybe the material basis of antioxidant fraction of P. lobata .

Objective: To analyze the expression of mismatch repair (MMR) protein and the EB virus infection in gastric adenocarcinoma, and to examine the association of MMR expression and EB virus infection with clinicopathological parameters. Methods: A case-control study was performed. Clinicopathological data of patients who was pathologically diagnosed as gastric adenocarcinoma, received radical gastrectomy and had complete clinicopathological data from August 2017 to April 2020 in Tianjin Medical University Cancer Institute and Hospital were retrospectively collected and analyzed. The immunohistochemistry (IHC) of MMR proteins and in situ hybridization (ISH) of Epstein-Barr virus encoded RNA (EBER) were reviewed. The associations of MMR and EBER results with clinicopathological parameters were analyzed. The main observations of the study were MMR and EBER expression, and association of MMR and EBER results with clinicopathological parameters. Results: Eight hundred and eighty-six patients were enrolled, including 98 patients who received preoperative neoadjuvant chemoradiotherapy. Of 886 patients, 613 (69.2%) were males and the median age was 60 (22-83) years; 831 (93.8%) were mismatch repair proficiency (pMMR), and 55 (6.2%) were mismatch repair deficiency (dMMR). In dMMR group, 47 cases (85.5%) had the deficiency of both MLH1 and PMS2, 1 case (1.8%) had the deficiency of both MSH2 and MSH6, 4 cases (7.3%) had the deficiency only in PMS2, 2 cases (3.6%) had the deficiency only in MSH6, and 1 case (1.8%) had the deficiency only in MSH2. The deficiency rates of PMS2, MLH1, MSH6 and MSH2 were 5.8% (51/886), 5.3% (47/886), 0.3% (3/886) and 0.2% (2/886), respectively. Among the 871 cases with EBER results, 4.9% (43/871) were positive EBER. Univariate analysis showed that dMMR was more frequently detected in female patients (χ(2)=10.962, P=0.001), cancer locating in the antrum (χ(2)=9.336,P=0.020), Lauren intestinal type (χ(2)=9.718, P=0.018), stage T3 (χ(2)=25.866, P<0.001) and TNM stage II (χ(2)=15.470, P=0.002). The ratio of dMMR was not significantly associated with age, tumor differentiation, histological type, lymph node metastasis, distant metastasis or Her-2 immunohistochemical score (all P>0.05). Compared with negative EBER, positive EBER was more frequent in male patients (χ(2)=9.701, P=0.002), cancer locating in gastric fundus and corpus (χ(2)=17.964, P<0.001), gastric cancer with lymphoid stroma (χ(2)=744.073, P<0.001) and poorly differentiated cancer (χ(2)=13.739, P=0.010). Positive EBER was not significantly associated with age, depth of invasion, lymph node metastasis, distant metastasis, TNM stage or Her-2 immunohistochemical score (all P>0.05). In addition, all dMMR cases were EBER negative, and all cases of positive EBER were pMMR. Conclusions: The positive EB virus status is mutually exclusive with dMMR, indicating that different molecular subtypes of gastric adenocarcinoma are involved in different molecular pathways in tumorigenesis and progression. The overlapping of dMMR or positive EBER status and positive Her-2 expression is found in some cases of gastric adenocarcinoma. Patients with gastric adenocarcinoma after radical surgery should be tested for MMR status if they are female, the tumor locates in gastric antrum, the TNM staging is stage II or T3, or if the Lauren classification is intestinal type. And if patients are male, the tumor locates in the gastric fundus and corpus, the cancer is lymphoid stroma, or poor differentiated, the expression of EBER should be detected. Results of our study may provide evidence for further decision-making of clinical treatment.
Adenocarcinoma ; Case-Control Studies ; DNA Mismatch Repair ; Epstein-Barr Virus Infections ; Female ; Herpesvirus 4, Human ; Humans ; Male ; Middle Aged ; Mismatch Repair Endonuclease PMS2/metabolism* ; MutL Protein Homolog 1/genetics* ; MutS Homolog 2 Protein/metabolism* ; Retrospective Studies ; Stomach Neoplasms