ObjectiveTo optimize and validate the optimal sequential alcohol-water extraction process of modified Banxia Xiexintang（MBXT） based on pharmacodynamic evaluation， combined with the G1-entropy weight method and Box-Behnken response surface methodology， and to systematically and comprehensively analyze the material basis of this formula， providing a scientific basis for its quality control and industrial production. MethodsRats were randomly divided into the normal group， model group， metformin group， and MBXT water extraction， water extraction and ethanol precipitation， sequential ethanol-water extraction groups. Except for the normal group， a polycystic ovary syndrome with insulin resistance（PCOS-IR） model was established in all rats via a high-fat diet combined with letrozole induction. Enzyme-linked immunosorbent assay（ELISA） biochemical assay kits and hematoxylin-eosin（HE） staining were used to compare sex hormone levels in serum and ovarian histopathology， thereby screening extraction process routes. Based on this， a comprehensive score was constructed using the G1-entropy weight method based on the transfer rates of index components（berberine hydrochloride and baicalin） and the dry extract rate. Box-Behnken response surface methodology was then utilized to optimize the extraction process parameters. Finally， the chemical constituents of the sample from the optimal process were qualitatively analyzed by ultra-performance liquid chromatography-quadrupole-electrostatic field orbitrap mass spectrometry（UPLC-Q-Orbitrap-MS）. ResultsPharmacodynamic findings revealed that compared with the normal group， serum testosterone（T） and luteinizing hormone（LH） levels were significantly elevated in the model group， while estradiol（E₂） and follicle-stimulating hormone（FSH） levels were significantly decreased（P<0.01）， with polycystic changes observed in ovarian tissues. Compared with the model group， all treatment groups significantly reversed the changes in sex hormone levels， with the sequential ethanol-water extraction group showing the optimal effect in improving the aforementioned indicators and pathological morphology， followed by subsequent process optimization. The optimized process involved adding 12 times the amount of 70% ethanol for extracting twice， each lasting 120 min， and adding 12 times the amount of water for extracting thrice， each lasting 90 min. Validation test results showed that under optimal process conditions， the average transfer rates of berberine hydrochloride and baicalin were 76.05% and 93.38%， respectively. MS analysis identified a total of 377 compounds， including 112 flavonoids， 41 terpenoids， 28 organic acids， 22 coumarins， and 8 alkaloids， while elucidating the cleavage patterns of key components. ConclusionThe optimized sequential ethanol-water extraction process is stable and feasible， effectively preserving the material basis of MBXT for treating PCOS-IR. It further clarifies the main chemical composition of this formula， providing a scientific basis for the development and quality control of its preparations.

ObjectiveTo evaluate the therapeutic effects of modified Banxia Xiexintang（MBXT） on polycystic ovary syndrome with insulin resistance（PCOS-IR） rats and reveal its potential mechanisms based on the integrated analysis of transcriptomics and metabolomics. MethodsFemale SD rats were selected， and a PCOS-IR model was established by intragastric administration of letrozole combined with a high-fat diet for 21 days. The modeled rats were randomly divided into the model group， MBXT low-， medium-， and high-dose groups（6.62， 13.23， 26.46 g·kg^-1）， and metformin group（0.158 g·kg^-1）， with a normal group set up separately. After 14 days of administration， the estrous cycle was observed， ovarian morphology was examined by hematoxylin-eosin（HE） staining， and the levels of testosterone（T）， estradiol（E₂）， follicle-stimulating hormone（FSH）， and luteinizing hormone（LH） in serum were detected by enzyme-linked immunosorbent assay（ELISA）. Serum metabolites and ovarian tissue gene expression were detected using ultra-performance liquid chromatography-quadrupole-electrostatic orbitrap mass spectrometry（UPLC-Q-Orbitrap-MS） and RNA-Seq technology， respectively， followed by multi-omics integrated analysis. ResultsPharmacodynamic findings revealed that all MBXT dose groups could reversed abnormal estrous cycles in PCOS-IR rats， improve polycystic ovarian lesions， and normalize dysregulated serum hormone levels（T， LH， E₂， FS， P<0.05， P<0.01）. Metabolomic analysis revealed that compared with the model group， MBXT reversed 278 differential metabolites such as estrone and S-formylglutathione， mainly involving pathways such as steroid hormone biosynthesis， glutathione metabolism， and lipid peroxidation regulation. Transcriptomic analysis identified 434 differentially expressed genes， and enrichment analysis revealed that MBXT significantly regulated lipid peroxidation defense systems， including glutathione metabolism， peroxisome function， and fatty acid metabolism， thereby intervening in ferroptosis processes. It also engaged in inflammation-related pathways such as the chemokine signaling pathway. Integrated analysis revealed that both metabolomics and transcriptomics co-enriched metabolic pathways associated with ferroptosis and fatty acid metabolism. And key Hub genes［such as Ras-related C3 botulinum toxin substrate 2 gene（Rac2） and Fas ligand gene（Faslg）］ showed significant correlations with differential metabolites. ConclusionMBXT can effectively ameliorate reproductive dysfunction and metabolic disorders in PCOS-IR rats. Its mechanism may be related to remodeling the immune-metabolism network， particularly by regulating MHC-mediated immune responses， inhibiting local ovarian ferroptosis， and enhancing steroid hormone synthesis pathways.

ObjectiveTo investigate the mechanism of modified Banxia Xiexintang（MBXT） in improving ovarian dysfunction in polycystic ovary syndrome with insulin resistance（PCOS-IR） rats by inhibiting ferroptosis through the adenosine monophosphate（AMP）-activated protein kinase（AMPK）/fatty acid synthase（FASN）/glutathione peroxidase 4（GPX4） signaling pathway. MethodsSeventy-six female SD rats were randomly divided into a normal group（n=13） and a modeling group（n=63）. The modeling group established a PCOS-IR model by intragastric administration of letrozole combined with a high-fat diet for 21 days. After successful modeling， these rats were randomly divided into the model group， MBXT low-， medium-， and high-dose groups（6.62， 13.23， 26.46 g·kg^-1）， metformin group（0.158 g·kg^-1）， and high-dose of MBXT combined with ferroptosis inducer Erastin group（15 mg·kg^-1）， with 10 rats in each group. After 14 days of intervention， ovarian pathological morphology was observed by hematoxylin-eosin（HE） staining， the mitochondrial ultrastructure of granulosa cells was observed by transmission electron microscopy（TEM）， ovarian reactive oxygen species（ROS） levels were detected by dihydroethidium（DHE） probe， biochemical methods were used to detect Fe²⁺， malondialdehyde（MDA）， glutathione（GSH） and other indicators in ovarian tissues， serum sex hormone and insulin levels were measured by enzyme-linked immunosorbent assay（ELISA）， and the protein expressions of AMPK， FASN， acyl-CoA synthetase long-chain family member 4（ACSL4）， GPX4， and solute carrier family 7 member 11（SLC7A11） in ovarian tissues were detected by Western blot. ResultsCompared with the normal group， the model group showed polycystic changes in the ovaries， with atrophy of mitochondria in granulosa cells and increased membrane density. Serum levels of testosterone（T）， luteinizing hormone（LH）， and insulin were significantly increased（P<0.01）. The levels of ROS， MDA， 4-hydroxynonenal（4-HNE）， and Fe²⁺ in ovarian tissues were significantly elevated（P<0.01）， while adenosine triphosphate（ATP）， GSH， and reduced nicotinamide adenine dinucleotide phosphate （NADPH） levels were significantly decreased（P<0.01）. The phosphorylation levels of AMPK and acetyl-CoA carboxylase （ACC）， as well as the protein expressions of SLC7A11， GPX4， and ferroptosis suppressor protein 1（FSP1） were significantly downregulated（P<0.01）， whereas the expressions of FASN， ACSL4， and nuclear receptor coactivator 4（NCOA4） were significantly upregulated（P<0.01）. Compared with the model group， MBXT intervention at various doses improved the above pathological changes and biochemical indicators in a dose-dependent manner， with the high-dose group showing the most significant effect（P<0.01）. Compared with the MBXT high-dose group， the high-dose of MBXT combined with ferroptosis inducer Erastin group restored ovarian ferroptosis characteristics in rats， with increased ROS and lipid peroxidation products， and altered expressions of key proteins（P<0.05， P<0.01）. ConclusionMBXT can effectively improve ovarian function and metabolic disorders in PCOS-IR rats. Its mechanism may be related to activating the AMPK/ACC signaling pathway， downregulating FASN and ACSL4 to reduce lipid peroxidation substrates， and restoring glucose-6-phosphate dehydrogenase/phosphoglycerate dehydrogenase（G6PD/PHGDH） metabolic flux to enhance the GPX4/FSP1 antioxidant defense system， thereby inhibiting ferroptosis in ovarian granulosa cells.

ObjectiveTo investigate the characteristics and distribution of traditional Chinese medicine （TCM） syndromes in the patients with esophageal squamous cell carcinoma （ESCC）. MethodsAn electronic questionnaire was developed to collect the general data and four examination information of ESCC patients treated in 10 areas with high incidence of esophageal cancer in China from June 2020 to March 2021. Multiple analyses including frequency analysis， factor analysis， and hierarchical cluster analysis were performed to analyze the potential syndrome elements， disease location， and common syndromes of ESCC. ResultsA total of 2 027 patients with ESCC were included. Statistical analysis was performed on 113 symptoms， physical signs， 33 tongue manifestation variables， and 23 pulse manifestation variables of the patients’ four examination information. Factor analysis was performed on 55 variables with frequency>10%， extracting 19 common factors. According to clinical experience and expert opinions， the main lesions of patients with ESCC were in the spleen and stomach， and the main syndrome elements were Qi stagnation， blood stasis， phlegm， dampness， and Qi deficiency， with the syndrome element combination of phlegm obstruction + Qi stagnation + blood stasis being the most common. The syndromes can be classified into four categories of liver-stomach disharmony + combined phlegm and Qi obstruction， kidney-spleen dysfunction + combined phlegm and stasis， spleen-kidney Yang deficiency + obstinate phlegm and blood stasis， and liver-kidney Yin deficiency + obstinate phlegm and blood stasis. The main syndrome of ESCC was liver-stomach disharmony + combined phlegm and Qi obstruction in the early stage， liver-spleen dysfunction + combined phlegm and stasis in the middle stage， and spleen-kidney Yang deficiency + obstinate phlegm and blood stasis in the late stage. ConclusionESCC mainly has main pathological features of internal deficiency and external excess and combined deficiency and excess， with the key syndrome elements being phlegm obstruction， Qi stagnation， and blood stasis. The main disease locations are in the spleen and stomach， involving the liver， kidney， chest and diaphragm， heart， and lung. The main syndrome is liver-stomach disharmony + combined phlegm and Qi obstruction. In clinical practice， it is necessary to grasp the pathogenesis dynamics of the disease and use prescriptions according to patients’ syndromes.

(±)-Talapyrones A-F (1-6), six pairs of dimeric polyketide enantiomers featuring unusual 6/6/6 and 6/6/6/5 ring systems, were isolated from the fungus Talaromyces adpressus. Their structures were determined by spectroscopic analysis and HR-ESI-MS data, and their absolute configurations were elucidated using a modified Mosher's method and electronic circular dichroism (ECD) calculations. (±)-Talapyrones A-F (1-6) possess a 6/6/6 tricyclic skeleton, presumably formed through a Michael addition reaction between one molecule of α-pyrone derivative and one molecule of C₈ poly-β-keto chain. In addition, compounds 2/3 and 4/5 are two pairs of C-18 epimers, respectively. Putative biosynthetic pathways of 1-6 were discussed.
Polyketides/isolation & purification* ; Talaromyces/chemistry* ; Stereoisomerism ; Molecular Structure ; Circular Dichroism ; Pyrones/chemistry*

Objective:To discuss the effect of circadion rhythm gene TIMELESS(TIM)silencing on immune escape of the ovarian cancer cells,and to clarify its related mechanism.Methods:The CD8+T lymphocytes were isolated and identified by flow cytometry to detect the proportion of CD3+/CD8+cell subsets.The human ovarian cancer SK-OV-3 cells were cultured in vitro and divided into interference plasmid transfected with TIM small interfering(siRNA)(si-TIM),negative control plasmid(si-NC),programmed death ligand 1(PD-L1)over-expression plasmid(oe-PD-L1),and negative control plasmid(oe-NC)groups.The cells were further divided into blank control group(BC group,non-transfection),si-NC group(transfected with si-NC),si-TIM group(transfected with si-TIM),si-NC+oe-NC group(transfected with si-NC and oe-NC),and si-TIM+oe-PD-L1 group(transfected with si-TIM and oe-PD-L1).Real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting methods were used to detect the expression levels of TIM mRNA and protein in the SK-OV-3 cells to verify TIM gene silencing.The transfected SK-OV-3 cells were co-cultured with activated CD8+T lymphocytes and divided into BC group(SK-OV-3 cells cultured alone),BC/T group,si-NC/T group,si-TIM/T group,si-NC+oe-NC/T group,and si-TIM+oe-PD-L1/T group.CCK-8 method was used to detect the survival rates of the SK-OV-3 cells in various groups;flow cytometry was used to detect the apoptotic rates of the SK-OV-3 cells and positive expression rate of PD-L1 on surface of the cells in various groups;enzyme-linked immunosorbent assay(ELISA)method was used to detect the levels of interferon-γ(IFN-γ)and tumor necrosis factor-α(TNF-α)in the co-culture supernatant;lactate dehydrogenase(LDH)release assay was used to detect the cytotoxicity of the CD8+T lymphocytes in various groups;RT-qPCR method was used to detect the expression levels of TIM and PD-L1 mRNA in the SK-OV-3 cells in various groups;Western blotting method was used to detect the expression levels of TIM and PD-L1 proteins in the SK-OV-3 cells in various groups.Results:After scparated with immune magnetic bead method,the proportion of CD8+T lymphocyte(CD3+/CD8+)subsets was(96.56%±0.59%),indicating high purity of the extracted CD8+T lymphocytes.Compared with BC group,the expression levels of TIM mRNA and protein in the cells in si-TIM group were significantly decreased(P<0.01),suggesting successful TIM gene silencing in the ovarian cancer SK-OV-3 cells.The CCK-8 results showed that compared with BC group,the survival rate of the SK-OV-3 cells in BC/T group was significantly decreased(P<0.01);compared with BC/T group,the survival rate of the SK-OV-3 cells in si-TIM/T group was significantly decreased(P<0.01).The flow cytometry results showed that compared with BC group,the apoptotic rate of the SK-OV-3 cells in BC/T group was significantly increased(P<0.01);compared with BC/T group,the apoptotic rate of the SK-OV-3 cells in si-TIM/T group was significantly increased(P<0.01);compared with si-TIM/T group,the apoptotic rate of the SK-OV-3 cells in si-TIM+oe-PD-L1/T group was significantly decreased(P<0.01).Compared with BC group,the positive expression rate of PD-L1 on surface of the SK-OV-3 cells in si-TIM group was significantly decreased(P<0.01).The ELISA results showed that compared with BC/T group,the levels of IFN-γ and TNF-α in the culture supernatant in si-TIM/T group were significantly increased(P<0.01);compared with si-TIM/T group,the levels of IFN-γ and TNF-α in the supernatant in si-TIM+oe-PD-L1/T group were significantly decreased(P<0.01).The LDH release assay results showed that compared with BC/T group,the cytotoxicity of the CD8+T lymphocytes in si-TIM/T group was significantly increased(P<0.01);compared with si-TIM/T group,the cytotoxicity of the CD8+T lymphocytes in si-TIM+oe-PD-L1/T group was significantly weakened(P<0.01).The RT-qPCR and Western blotting results showed that compared with BC group,the expression levels of PD-L1 mRNA and protein in the SK-OV-3 cells in si-TIM group were significantly decreased(P<0.01);compared with si-TIM group,the expression level of PD-L1 protein in the cells in si-TIM+oe-PD-L1 group was significantly increased(P<0.01).Conclusion:TIM gene silencing enhances the cytotoxic effect of CD8+T lymphocytes on ovarian cancer SK-OV-3 cells and inhibits immune escape,and its mechanism may be related to the regulation of PD-L1 protein expression.

Objective:To observe the efficacy and safety of combined lienal polypeptide injection therapy in the treatment of Mycoplasma pneumoniae pneumonia（MPP）in children aged 3 to 14 years old in multiple clinical centers.Methods:A randomized，controlled，multi-center clinical study design was adopted.A total of 240 hospitalized children aged 3 to 14 years old with MPP from 7 hospitals from September 1，2023 to January 31，2024 were included.According to the severity of pneumonia，they were divided into the mild MPP group with 80 cases and the severe MPP/refractory MPP（SMPP/RMPP）group with 160 cases，and then randomly divided into the control group and the experimental group at a ratio of 1 ∶1，using the random number table method.After screening，subjects entered a treatment period of 5 to 7 days.The control group was treated with azithromycin，while the experimental group was treated with azithromycin plus lienal polypeptide injection .The recovery of lung CT，length of hospital stay，duration of fever，cough score，whether mild cases developed into severe or refractory cases，duration of hormone use，use of intravenous immunoglobulin（IVIG），bronchoscopy treatment，and immune function were observed between the two groups to evaluate the efficacy of lienal polypeptide injection.Adverse events after medication，vital signs，blood routine，urine routine，liver function，myocardial enzymes，renal function，and electrocardiogram were observed to evaluate the safety. Results:A total of 231 subjects have completed the trial in the 7 hospitals，including 118 cases in the experimental group and 113 cases in the control group.Main observation index：the rate of lung CT aggravation in the experimental group was lower than that in the control group（2.6% vs 15.3%， P<0.01），and the difference was statistically significant.Secondary indexes：there were no statistically significant differences in the length of hospital stay，duration of fever，cough score，duration of hormone use，whether IVIG treatment was used，the number of bronchoscopy treatment cases，and immunoglobulin between the two groups（all P>0.05）.However，the rate of cases of plastic bronchitis（PB）found under bronchoscopy in the experimental group was lower than that in the control group（0 vs 18.8%， P=0.03），and the difference was statistically significant.Among the mild MPP（72 cases），there were no statistically significant differences in the length of hospital stay，duration of fever，cough score，duration of hormone use，whether IVIG treatment was used，the number of bronchoscopy treatment cases，and the improvement rate of lung CT between the two groups（all P>0.05）.However，compared with the control group，the rate of cases developing into SMPP/RMPP in the experimental group was less（24.3% vs 48.6%， P=0.03），and the difference in IgG before and after treatment was small［0.53（-0.04，1.18）g/L vs 1.33（0.48，2.25）g/L， P=0.01］.Among the SMPP/RMPP cases（159 cases），the rate of cases of PB found under bronchoscopy in the experimental group was less than that in the control group（0 vs 20%， P=0.04），and the rate of cases with aggravated lung CT in the experimental group was less than that in the control group（1.3% vs 19.5%， P<0.01），and the improvement rate of lung CT in the experimental group was higher than that in the control group（88.8% vs 75.3%， P=0.03），with statistically significant differences.There were no statistically significant differences in the length of hospital stay，duration of fever，cough score，duration of hormone use，whether IVIG treatment was used，the number of bronchoscopy treatment cases，and immunoglobulin between the two groups（all P>0.05）.Two cases in the experimental group developed rashes，which improved after the drug was discontinued.There were no serious adverse reactions such as abnormal vital signs like dyspnea and cyanosis due to the use of lienal polypeptide injection.There were no obvious changes in blood routine，liver function，myocardial enzymes，renal function，electrocardiogram，and urine routine values before and after medication compared with the baseline. Conclusion:The combined use of lienal polypeptide injection in the treatment of MPP in children can reduce the probability of the transformation from mild cases to SMPP/RMPP，reduce the rate of aggravation of the image findings，promote the absorption of lung inflammation，reduce the rate of PB found under bronchoscopy，and has good safety.

Da Chaihutang from the Treatise on Febrile Disease （Shanghanlun） has the function of harmonizing lesser Yang and discharging internal heat. It is formulated by ZHANG Zhongjing for the concurrent diseases of lesser Yang and Yang brightness and has been widely used in the treatment of digestive system diseases， especially malignant tumors. By review of the articles published in the last 20 years， this paper summarizes the application of Da Chaihutang in treating digestive system tumors from syndrome analysis， clinical research， and mechanism research. Da Chaihutang can treat the syndrome involving lesser Yang and Yang brightness in the digestive system， release interior and exterior to expel pathogen， and remove obstruction by conforming to the descending nature of the six fu-organs. In clinical practice， Da Chaihutang can directly treat digestive system malignant tumors such as liver cancer， pancreatic cancer， intestinal cancer， gastric cancer， and gallbladder cancer. In addition， it can relieve common complications of digestive system malignant tumors， such as cancerous fever， malignant obstructive jaundice， and constipation. Moreover， it can alleviate the adverse reactions caused by Western medical treatment， such as post-embolization syndrome， side effects of chemotherapy， and incomplete postoperative obstruction. Da Chaihutang is effective when used alone as it can relieve clinical symptoms， improve prognosis， and prolong survival of advanced patients and is safe and non-toxic， suitable for long-term use by tumor patients. Regarding the mechanism， Da Chaihutang can promote the apoptosis and inhibit the proliferation of tumor cells， reduce inflammation and inflammatory injury， and improve the liver function. The clear effect and mechanism confirms the anti-tumor effect of Da Chaihutang. This paper comprehensively describes the current research status of Da Chaihutang in the treatment of digestive system tumors and puts forward the deficiencies and improvement measures for the current research， aiming to provide reference for the application of this formula in treating digestive system tumors， the establishment of Chinese and Western medicine treatment schemes of tumors， and the research and development of anti-tumor drugs.

In March 2020, the outflow of age limited videos from ICU in Spain inspired us to rethink whether there is age discrimination in the allocation of scarce medical resources. This paper frist reflected on the problem of age discrimination caused by this phenomenon from four moral intuitions: the sacred view of life, the quality of life and values, public health ethics and Chinese culture, and then examined whether it is illegal from the legal level, finally pointed out the negative impact on the society, and put forward that taking age as the standard for the allocation of scarce medical resources is not suitable for China’s national conditions.

BACKGROUND:Numerous clinical studies have suggested a close relationship between obesity and osteoporosis,but whether there is a genetic causal effect between obesity and osteoporosis remains unclear. OBJECTIVE:To explore the association between obesity and osteoporosis using summary data from a large-scale genome-wide association study(GWAS)through Mendelian randomization analysis. METHODS:Obesity data were derived from summary statistics of the Genetic Investigation of Anthropometric Traits(GIANT)and the UK Biobank(UKBB).Osteoporosis data were obtained from the Genetic Factors for Osteoporosis(GeFOS)consortium,including two bone density phenotypes:total body bone mineral density(BMD)and heel BMD.The inverse variance-weighted method was the primary analysis,with the Mendelian randomization method based on Egger regression(MR-Egger)and weighted median method as supplementary approaches to calculate the causal association between genetic variations related to obesity and osteoporosis.Sensitivity analyses were conducted to validate the reliability of the results.Heterogeneity was assessed using Cochran's Q test.Horizontal pleiotropy was assessed through the MR-Egger intercept test.Leave-one-out analysis was performed to evaluate the potential influence of single nucleotide polymorphisms on the combined inverse variance-weighted estimates. RESULTS AND CONCLUSION:(1)Impact of obesity on osteoporosis:In addition to body mass index and forearm BMD,body mass index,waist-to-hip ratio,body mass index-adjusted waist-to-hip ratio,and whole-body body mass index,heel BMD,forearm BMD,lumbar spine BMD,and femoral neck BMD were causally related to each other.Further Meta-analysis revealed that obesity increased the risk of BMD(odds ratio=1.07,95%confidence interval:1.03-1.12,P<0.01).(2)Impact of osteoporosis on obesity:Apart from arm BMD and lumbar spine BMD as exposure factors showing causal relationships with obesity,other datasets indicated no causal effect between total body BMD,heel BMD,femoral neck BMD,and obesity.Additional meta-analysis demonstrated that BMD did not increase the risk of obesity(odds rate=0.99,95%confidence interval:0.98-1.01,P<0.01).There is a causal relationship between obesity and osteoporosis,suggesting that obesity may be a risk factor for osteoporosis.However,no causal association is found between osteoporosis and obesity.