OBJECTIVE: To observe the effects of Xingnao Kaiqiao acupuncture technique (for regaining consciousness and opening orifice) on methylation of N6-methyladenosine (m6A), and key methyltransferases and demethylases, so as to clarify the mechanism of acupuncture on cerebral ischemia-reperfusion injury (CIRI). METHODS: Of 68 male Sprague-Dawley rats of SPF grade, 15 rats were randomly selected as a sham-operation group, and the remaining rats were subjected to the model of middle cerebral artery occlusion using the suture ligation. CIRI was induced by ischemia for 2 h followed by reperfusion. Rats that failed to modeling or died were excluded. The rest 45 rats were randomly divided into three groups, i.e. model group, acupuncture group, and non-point acupuncture group, with 15 rats in each group. The rats in the acupuncture group were treated with acupuncture at bilateral "Neiguan" (PC6) and "Shuigou" (GV26). In the non-point acupuncture group, acupuncture was delivered at three non-points, located 3 mm below the bilateral midaxillary line and 3 mm lateral to the tip of the coccyx. One intervention was operated in these two acupuncture groups and the needles were retained for 30 min. Before modeling start and 2 h after ischemia, a laser speckle flowmeter was used to monitor the cerebral blood perfusion. In 2 h of ischemia and 24 h of reperfusion, the neurological behavioral score was evaluated. The volume of rat cerebral infarction was determined by triphenyltetrazolium chloride (TTC) staining, and the level of m6A methylation in ischemic cortical area was detected by Dot blot, and the protein and mRNA expression of the demethylase i.e. fat mass and obesity-associated protein (FTO), AlkB homolog 5 (ALKBH5) and key methyltransferases, i.e. methyltransferase-like 3 (METTL3), methyltransferase-like 14 (METTL14), and Wilms' tumor 1-associated protein (WTAP) in ischemic cortical area were detected by Western blot and real-time PCR. RESULTS: Cerebral blood perfusion decreased by>70% after 2 h ischemia. Compared with the sham-operation group, the neurobehavioral score and the percentage of cerebral infarction volume increased in the model group (P<0.01); the level of m6A methylation in the ischemic cortical area increased (P<0.01), the protein and mRNA expression of FTO decreased (P<0.01), and that of ALKBH5, METTL3, and METTL14 increased (P<0.01, P<0.05) in the model group. When compared with the model group and the non-point acupuncture group, the acupuncture group showed a decrease in the neurobehavioral score and the percentage of cerebral infarction volume (P<0.01), the level of m6A methylation in the ischemic cortical area was reduced (P<0.01, P<0.05), and the protein and mRNA expression of FTO was elevated (P<0.01). CONCLUSION Xingnao Kaiqiao acupuncture technique presents its protective effect on the brain in the rats with CIRI, which is related to up-regulating the expression of FTO and modulating m6A methylation.
Animals ; Rats, Sprague-Dawley ; Male ; Acupuncture Therapy ; Reperfusion Injury/genetics* ; Rats ; Brain Ischemia/genetics* ; Humans ; Adenosine/metabolism* ; Methylation ; Acupuncture Points ; Cerebral Cortex/metabolism*

OBJECTIVE: To observe the clinical efficacy of acupoint thread-embedding therapy of regulating governor vessel, dispersing lung, and suppressing reflux for gastroesophageal reflux cough (GERC). METHODS: A total of 120 GERC patients were randomly assigned to an observation group (60 cases, 1 case dropped out) and a control group (60 cases, 1 case was eliminated). The observation group received acupoint thread-embedding treatment at positive response points of governor vessel. If no such points were detected, the following acupoints were used: Dazhui (GV14), Fenghu (Extra), Shendao (GV11), Lingtai (GV10), and Zhiyang (GV9). Treatment was administered once every two weeks. The control group received oral rabeprazole enteric capsules at 20 mg twice daily. All the treatment was given for 6 weeks. Clinical outcomes were assessed using cough symptom score, reflux disease questionnaire (RDQ) score, and Leicester cough questionnaire (LCQ) score before and after treatment in the two groups. Clinical efficacy was also compared between the two groups. RESULTS: After treatment, both groups showed decreased cough symptom scores and the each item scores and total scores of RDQ (P<0.001), and increased LCQ scores (P<0.001) compare with those before treatment. The observation group exhibited lower cough symptom score and chest pain, reflux and total score of RDQ, and higher LCQ score compared to those in the control group (P<0.05). The total effective rate in the observation group was 94.9% (56/59), which was higher than 84.7% (50/59) in the control group (P<0.05). CONCLUSION Acupoint thread-embedding therapy of regulating governor vessel, dispersing lung, and suppressing reflux could effectively alleviate cough and reflux symptoms in patients with GERC and improve their quality of life.
Humans ; Acupuncture Points ; Gastroesophageal Reflux/physiopathology* ; Male ; Female ; Cough/physiopathology* ; Middle Aged ; Aged ; Acupuncture Therapy ; Adult ; Treatment Outcome ; Lung/physiopathology* ; Meridians

OBJECTIVES: To examine the changes in serum levels of endoplasmic reticulum stress (ERS) proteins GRP78/CHOP in patients with lupus nephritis (LN) and analyze their diagnostic value and association with renal pathological features. METHODS: From a sample bank established based on a multicenter cohort study of systemic lupus erythematosus (SLE), 60 LN patients and 35 SLE patients without renal involvement were randomly selected. ELISA was used to detect serum levels of GRP78 and CHOP in the patients to analyze their correlation with clinical features and their diagnostic ability for LN and active LN. MRL/lpr mice were used as an animal model of LN to examine their serum levels of GRP78 and CHOP expression and renal expressions of endoplasmic reticulum apoptosis-related proteins. RESULTS: Serum GRP78 and CHOP levels were significantly higher in LN patients than in SLE patients without renal involvement (P<0.05), and were also higher in active LN patients than in patients in the stable phase (P<0.05). Correlation analysis indicated that serum GRP78 and CHOP levels were positively correlated with SLEDAI scores and 24-h urinary protein. ROC analysis showed that CHOP had a high diagnostic ability for LN (AUC=0.762) and active LN (AUC=0.933). Consistent with the clinical findings, serum GRP78 and CHOP levels were elevated in LN mice, and the expressions of PERK and IRE1α pathway proteins were also increased in the kidneys of the mice. TUNEL staining showed increased renal cell apoptosis and elevated renal expressions of apoptosis-related proteins in LN mice. CONCLUSIONS Serum levels of GRP78/CHOP are increased in LN patients possibly in association with ERS-induced apoptosis mediated by the PERK/IRE1α dual pathway.
Endoplasmic Reticulum Chaperone BiP ; Lupus Nephritis/blood* ; Transcription Factor CHOP/blood* ; Heat-Shock Proteins/blood* ; Animals ; Apoptosis ; Humans ; Mice ; Mice, Inbred MRL lpr ; Female ; Adult ; Endoribonucleases/metabolism* ; Male ; eIF-2 Kinase/metabolism* ; Protein Serine-Threonine Kinases/metabolism* ; Young Adult ; Endoplasmic Reticulum Stress ; Kidney/metabolism* ; Middle Aged ; Signal Transduction

Sjögren's syndrome (SS) is an autoimmune disease characterized primarily by oral and periocular dryness. Astragalus-Salvia (AS) and Ophiopogon-Dendrobium (OD) represent two frequently utilized herb pairs in SS treatment. While the combination of AS-OD herb pairs demonstrates clinical efficacy in alleviating SS symptoms, its underlying mechanism remains unclear. This investigation sought to assess the therapeutic effects and elucidate the potential mechanisms of AS-OD in non-obese diabetic (NOD)/Ltj mice with SS. The study utilized NOD/Ltj mice as SS models, administering AS-OD treatment for 10 weeks at doses of 113.1, 226.2, and 339.3 mg·d^-1·20 g^-1. Results demonstrated that AS-OD improved SS symptoms, evidenced by enhanced salivary flow rate, decreased anti-SSA/Ro and anti-SSB/La antibody levels, increased swimming duration, and reduced lactate (LA) and blood urea nitrogen (BUN) levels in NOD/Ltj mice. AS-OD reduced lymphocyte infiltration, enhanced Aquaporin-5 (AQP5) expression in the submandibular gland, decreased inflammatory cytokine levels in the submandibular gland, and reduced the T helper type 17/regulatory T lymphocyte (Th17/Treg) cell ratio in the spleen. Transcriptomic and proteomic analyses indicated AS-OD's involvement in regulating phosphatidylinositol-3-kinase/protein kinase B (PI3K/AKT) and Janus kinase 3/signal transducer and activator of transcription 3 (JAK1/STAT3) pathways, with inhibitory effects validated in both NOD/Ltj mice submandibular gland and A-253 cells. Furthermore, AS-OD enhanced cell viability and reduced A-253 cell apoptosis through the PI3K/AKT pathway. In A-253 cells, AS-OD reduced inflammatory cytokine levels, CXC chemokine ligand 9/10 (CXCL9/10), and T-cell chemotaxis by inhibiting the JAK1/STAT3 pathway. AS-OD mitigates SS by suppressing inflammation and immune responses through the PI3K/AKT and JAK1/STAT3 pathways.
Animals ; STAT3 Transcription Factor/genetics* ; Sjogren's Syndrome/immunology* ; Mice, Inbred NOD ; Proto-Oncogene Proteins c-akt/genetics* ; Phosphatidylinositol 3-Kinases/genetics* ; Mice ; Drugs, Chinese Herbal/administration & dosage* ; Signal Transduction/drug effects* ; Janus Kinase 1/genetics* ; Humans ; Female ; Astragalus Plant/chemistry* ; Male

Lipophagy, the selective engulfment of lipid droplets (LDs) by autophagosomes for lysosomal degradation, is critical to lipid and energy homeostasis. Here we show that the lipid transfer protein ORP8 is located on LDs and mediates the encapsulation of LDs by autophagosomal membranes. This function of ORP8 is independent of its lipid transporter activity and is achieved through direct interaction with phagophore-anchored LC3/GABARAPs. Upon lipophagy induction, ORP8 has increased localization on LDs and is phosphorylated by AMPK, thereby enhancing its affinity for LC3/GABARAPs. Deletion of ORP8 or interruption of ORP8-LC3/GABARAP interaction results in accumulation of LDs and increased intracellular triglyceride. Overexpression of ORP8 alleviates LD and triglyceride deposition in the liver of ob/ob mice, and Osbpl8-/- mice exhibit liver lipid clearance defects. Our results suggest that ORP8 is a lipophagy receptor that plays a key role in cellular lipid metabolism.
Animals ; Mice ; Lipid Droplets ; Autophagy ; Autophagosomes ; Homeostasis ; Triglycerides

AIM: To evaluate the clinical efficacy and safety of He-wei-zhi-xie (HWZX) capsules in diarrhea patients. METHODS: The clinical study was conducted in 35 clinical trials centers from October 2015 to December 2017 by multicenter, prospective, open and uncontrolled design methods. The primary efficacy endpoint is the effective rate of diarrhea, the secondary endpoints include recovery rate of diarrhea, recovery time of diarrhea, number of irregular stools and Leeds dyspepsia questionnaire. The pharmacodynamics model of time course was established by nonlinear mixed effect model, and the effect of covariates on pharmacodynamic parameters was investigated. The safety measures were the incidence of adverse events, adverse reactions and the laboratory test indicators. RESULTS: A total of 2 285 cases were included in full analysis set. The effective rate of diarrhea was 90.8%, and the diarrhea recovery rate was 77.3%. The median time of recovery was 3 days, and the Leeds score was reduced by 3.6 points. It is found that baseline has a significant effect on model parameter E

Postzygotic mutations are acquired in normal tissues throughout an individual's lifetime and hold clues for identifying mutagenic factors.Here,we investigated postzygotic mutation spectra of healthy individuals using optimized ultra-deep exome sequencing of the time-series samples from the same volunteer as well as the samples from different individuals.In blood,sperm,and muscle cells,we resolved three common types of mutational signatures.Signatures A and B represent clock-like mutational processes,and the polymorphisms of epigenetic regulation genes influence the pro-portion of signature B in mutation profiles.Notably,signature C,characterized by C＞T transitions at GpCpN sites,tends to be a feature of diverse normal tissues.Mutations of this type are likely to occur early during embryonic development,supported by their relatively high allelic frequencies,presence in multiple tissues,and decrease in occurrence with age.Almost none of the public datasets for tumors feature this signature,except for 19.6％of samples of clear cell renal cell carcinoma with increased activation of the hypoxia-inducible factor 1(HIF-1)signaling pathway.Moreover,the accumulation of signature C in the mutation profile was accelerated in a human embryonic stem cell line with drug-induced activation of HIF-1α.Thus,embryonic hypoxia may explain this novel signature across multiple normal tissues.Our study suggests that hypoxic condition in an early stage of embryonic development is a crucial factor inducing C＞T transitions at GpCpN sites;and indi-viduals'genetic background may also influence their postzygotic mutation profiles.

Objective To investigate the expression of serum pentraxin3（PTX3） and microRNA-199a（miR-199a） in patients with acute stroke，and to further analyze the relationship between PTX3 and miR-199a and the neurological function of the patients and their value in the diagnosis of the disease.Methods 83 patients with acute stroke who were treated in our hospital from January 2018 to December 2019 were selected as the study group. According to National Institute of Health stroke scale（NIHSS） score，the patients were divided into mild group（26 cases），moderate group（30 cases） and severe group（27 cases）.Another 40 volunteers who had physical examination in our hospital at the same time were selected as the control group. Serum levels of PTX3，miR-199a，soluble protein 100β（S100-β），and neuron specific enolase（NSE） in all subjects were detected.Results The levels of PTX3，S100-β and NSE in the study group were higher than those in the control group，and the levels of miR-199a in the study group were lower than those in the control group（P<0.05）；The levels of PTX3，S100 -β，NSE，NIHSS and miR-199a in severe group were higher than those in moderate group and mild group（P<0.05），and the levels of PTX3，S100-β，NSE and NIHSS in the moderate group were higher than those in the mild group，and the level of miR-199a was lower than that in the mild group（P<0.05）；Pearson analysis showed that PTX3 was positively correlated with S100-β，NSE and NIHSS scores in patients with acute stroke（P<0.05），and miR-199a was negatively correlated with S100-β，NSE and NIHSS scores（P<0.05）；ROC analysis showed that serum PTX3 and miR-199a had certain diagnostic value for acute stroke.The area under the curve was 0.864（95%CI 0.799～0.928）and 0.807（95%CI：0.704～0.910），respectively. The combination of the two could further improve the diagnostic value for acute stroke.Conclusion Serum PTX3 is highly expressed and miR-199a is lowly expressed in patients with acute stroke，and the expression of both is closely related to the patient’s nerve function.Both of them have a certain value of diagnosis.

The Genome Warehouse (GWH) is a public repository housing genome assembly data for a wide range of species and delivering a series of web services for genome data submission, storage, release, and sharing. As one of the core resources in the National Genomics Data Center (NGDC), part of the China National Center for Bioinformation (CNCB;https://ngdc.cncb.ac.cn), GWH accepts both full and partial (chloroplast, mitochondrion, and plasmid) genome sequences with different assembly levels, as well as an update of existing genome assemblies. For each assembly, GWH collects detailed genome-related metadata of biological project, biological sample, and genome assembly, in addition to genome sequence and annotation. To archive high-quality genome sequences and annotations, GWH is equipped with a uniform and standardized procedure for quality control. Besides basic browse and search functionalities, all released genome sequences and annotations can be visualized with JBrowse. By May 21, 2021, GWH has received 19,124 direct submissions covering a diversity of 1108 species and has released 8772 of them. Collectively, GWH serves as an important resource for genome-scale data management and provides free and publicly accessible data to support research activities throughout the world. GWH is publicly accessible at https://ngdc.cncb.ac.cn/gwh.

Objective:To explore the methods, complications and follow-up results of limb salvage for primary malignant tumors of the femoral shaft.Methods:From October 2006 to October 2019, 41 cases of primary malignant tumors of femoral shaft were analyzed retrospectively, 37 cases were followed up, including 22 males and 15 females, aged 8-46 years with an average age of 20.56±4.72 years old. All the lesions were located in the femoral shaft, and The Enneking stage was IIB. Tumor lesion ranged in the femur from 10 cm to 24 cm, and there was no pathological fracture. Pre-operative puncture biopsy was performed in all cases.22 cases were confirmed as osteosarcoma, 8 as chondrosarcoma, 5 as Ewing sarcoma, 2 as malignant fibrous histiocytoma. Neoadjuvant chemotherapy has been used to treat both osteosarcoma and Ewing sarcoma. Among the 37 patients, 8 cases of patients received total femoral prosthesis replacement, 14 cases of intramedullary nail fixation with allogenic bone graft, 8 cases of allograft-prosthetic composite (APC), and 7 cases of 3D printed prosthesis. The limb function was graded according to MSTS 93 system.Results:37 out of 41 patients were followed up, and the average follow-up time was 23.15±16.74 months (6-62 months). There were 9 patients with pulmonary metastases (26.47%), among which 6 patients passed away due to multiple metastases. 28 patients survived without tumor at last follow-up. The MSTS score was18-28 (average of 22.55±2.57). The average score of 3D printing prosthesis group was 27±1.74, allogeneic bone transplantation group was 22.85±2.59, total femoral prosthesis replacement group was 20.25±2.25, and the APC prosthesis group was 20.5±2.07. There was statistical difference between 3D printing group and other groups. The overall excellent and good rate of lower limb function was 79.41%, in which the 3D group was 100%, allogeneic bone group was 78.5%, APC prosthesis group was 87.5%, total femoral prosthesis replacement group was 62.5%. There was no statistical difference in the excellent and good rate of limb function among the groups. There weren't any cases of hip joint dislocation after total femoral prosthesis replacement. Delayed union of bone healing was seen in 3 cases of allogeneic bone transplantation and 2 cases APC patients. One patient suffered from the postoperative hemorrhage-related diseases-sciatic nerve compression, and the nerve recovered after emergency debridement. One patient suffered from poor wound healing. The overall complication rate was 24.32% (9/37).Conclusion:The patients with primary malignant tumors of the femoral shaft can effectively recover the lower extremity function by choosing the appropriate surgical scheme, in order to achieve the purpose of limb preservation and improve the quality of life of the patients. 3D printing prosthesis provides a new choice in the treatment of femoral shaft tumors.