Oral squamous cell carcinoma (OSCC) is a common head and neck malignancy. Approximately 50% to 60% of patients with OSCC are diagnosed at a locally advanced stage (clinical staging III-IVa). Even with comprehensive and sequential treatment primarily based on surgery, the 5-year overall survival rate remains below 50%, and patients often suffer from postoperative functional impairments such as difficulties with speaking and swallowing. Programmed death receptor-1 (PD-1) inhibitors are increasingly used in the neoadjuvant treatment of locally advanced OSCC and have shown encouraging efficacy. However, clinical practice still faces key challenges, including the definition of indications, optimization of combination regimens, and standards for efficacy evaluation. Based on the latest research advances worldwide and the clinical experience of the expert group, this expert consensus systematically evaluates the application of PD-1 inhibitors in the neoadjuvant treatment of locally advanced OSCC, covering combination strategies, treatment cycles and surgical timing, efficacy assessment, use of biomarkers, management of special populations and immune related adverse events, principles for immunotherapy rechallenge, and function preservation strategies. After multiple rounds of panel discussion and through anonymous voting using the Delphi method, the following consensus statements have been formulated: 1) Neoadjuvant therapy with PD-1 inhibitors can be used preoperatively in patients with locally advanced OSCC. The preferred regimen is a PD-1 inhibitor combined with platinum based chemotherapy, administered for 2-3 cycles. 2) During the efficacy evaluation of neoadjuvant therapy, radiographic assessment should follow the dual criteria of Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1 and immune RECIST (iRECIST). After surgery, systematic pathological evaluation of both the primary lesion and regional lymph nodes is required. For combination chemotherapy regimens, PD-L1 expression and combined positive score need not be used as mandatory inclusion or exclusion criteria. 3) For special populations such as the elderly (≥ 70 years), individuals with stable HIV viral load, and carriers of chronic HBV/HCV, PD-1 inhibitors may be used cautiously under the guidance of a multidisciplinary team (MDT), with close monitoring for adverse events. 4) For patients with a poor response to neoadjuvant therapy, continuation of the original treatment regimen is not recommended; the subsequent treatment plan should be adjusted promptly after MDT assessment. Organ transplant recipients and patients with active autoimmune diseases are not recommended to receive neoadjuvant PD-1 inhibitor therapy due to the high risk of immune related activation. Rechallenge is generally not advised for patients who have experienced high risk immune related adverse events such as immune mediated myocarditis, neurotoxicity, or pneumonitis. 5) For patients with a good pathological response, individualized de escalation surgery and function preservation strategies can be explored. This consensus aims to promote the standardized, safe, and precise application of neoadjuvant PD-1 inhibitor strategies in the management of locally advanced OSCC patients.

OBJECTIVE: To investigate the characteristics of Vitamin D (VitD) and bone metabolism in patients with knee osteoarthritis (KOA) concurrent with osteoporosis (OP). METHODS: A retrospective analysis was performed on 240 patients who were admitted to the orthopedics department between March 2019 and March 2024. Patients were stratified into four distinct groups according to their respective disease categories.There were 90 patients in the simple KOA group, comprising 13 males and 77 females, age ranged from 50 to 91 years old with an average of (68.48±8.96) years old. There were 90 patients in the simple OP group, comprising 7 males and 83 females, age ranged from 52 to 88 years old with an average of (69.60±8.94 )years old. There were 30 patients in the KOA with OP group, comprising 1 male and 29 females, age ranged from 51 to 91 years old with an average of(69.03±7.93) years old. There were 30 patients in the physical examination group, comprising 5 males and 25 females, age ranged from 53 to 79 years old with an average of(64.93±6.51) years old. The general data and the levels of osteocalcin (OC), β-CrossLaps, parathyroid hormone(PTH) and VitD in each group were observed. RESULTS: The level of VitD in KOA with OP group (19.62±10.38) ng·ml^-1 and OP group (20.65±10.50) ng·ml^-1 was lower than that in physical examination group (27.46±8.00) ng·ml^-1 and KOA group (24.01±9.11) ng·ml^-1 (P<0.05). There were significant differences in β- CrossLaps and PTH levels among the four groups (P<0.001, P=0.019, respectively), while there was no significant difference in OC levels (P=0.763). Compared with the two simple disease groups, the KOA with OP group had higher levels of β - CrossLaps(0.81±0.30) ng·ml^-1 (P<0.001). There were significant differences in β-CrossLaps and PTH between the simple KOA group(0.54±0.22) ng·ml^-1, (46.03±18.08) pg·ml^-1 and the physical examination group (0.44±0.19) ng·ml^-1, (36.65±9.63) pg·mL^-1(P=0.038;P=0.006). There was a significant difference in PTH between the OP group(43.85±14.30) ng·ml-1, and the physical examination group, P=0.004. There was a significant difference in Kallgren-Lawrence grading between KOA with OP group and KOA group (P=0.006). Within KOA with OP group, the differences of β-CrossLaps and VitD levels among different K-L grades were statistically significant (P=0.016). The level of OC, β-CrossLaps and PTH within KOA with OP group was significantly different at different VitD levels (P=0.013, P=0.033, P=0.046). CONCLUSION Patients with KOA complicated by OP exhibit greater disturbances in bone metabolism and reduced VitD levels, particularly reflected by elevated β-CrossLaps. These findings underscore the importance of early monitoring of bone turnover and VitD supplementation in advanced-stage KOA with bone loss.
Humans ; Female ; Male ; Middle Aged ; Aged ; Vitamin D/blood* ; Osteoporosis/complications* ; Aged, 80 and over ; Osteoarthritis, Knee/complications* ; Retrospective Studies ; Bone and Bones/metabolism* ; Parathyroid Hormone/metabolism* ; Osteocalcin/metabolism*

ObjectiveTo explore the syndrome elements and evolving patterns of patients with hepatitis B virus-related acute on chronic liver failure （HBV-ACLF） at different stages. MethodsClinical information of 1,058 hospitalized HBV-ACLF patients， including 618 in the early stage， 355 in the middle stage， and 85 in the late stage， were collected from 18 clinical centers across 12 regions nationwide from January 1， 2012 to February 28， 2015. The “Hepatitis B-related Chronic and Acute Liver Failure Chinese Medicine Clinical Questionnaire” were designed to investigate the basic information of the patients， like the four diagnostic information （including symptoms， tongue， pulse） of traditional Chinese medicine （TCM）， and to count the frequency of the appearance of the four diagnostic information. Factor analysis and cluster analysis were employed to determine and statistically analyze the syndrome elements and patterns of HBV-ACLF patients at different stages. ResultsThere were 76 four diagnostic information from 1058 HBV-ACLF patients， and 53 four diagnostic information with a frequency of occurrence ≥ 5% were used as factor analysis entries， including 36 symptom information， 12 tongue information， and 5 pulse information. Four types of TCM patterns were identified in HBV-ACLF， which were liver-gallbladder damp-heat pattern， qi deficiency and blood stasis pattern， liver-kidney yin deficiency pattern， and spleen-kidney yang-deficiency pattern. In the early stage， heat （39.4%， 359/912） and dampness （27.5%， 251/912） were most common， and the pattern of the disease was dominated by liver-gallbladder damp-heat pattern （74.6%， 461/618）； in the middle stage， dampness （30.2%， 187/619） and blood stasis （20.7%， 128/619） were most common， and the patterns of the disease were dominated by liver-gallbladder damp-heat pattern （53.2%， 189/355）， and qi deficiency and blood stasis pattern （27.6%， 98/355）； and in the late stage， the pattern of the disease was dominated by qi deficiency （26.3%， 40/152） and yin deficiency （20.4%， 31/152）， and the patterns were dominated by qi deficiency and blood stasis pattern （36.5%， 31/85）， and liver-gallbladder damp-heat pattern （25.9%， 22/85）. ConclusionThere are significant differences in the distribution of syndrome elements and patterns at different stages of HBV-ACLF， presenting an overall trend of evolving patterns as "from excess to deficiency， transforming from excess to deficiency"， which is damp-heat → blood stasis → qi-blood yin-yang deficiency.

This study explores the effects and possible mechanisms of nuclear factor E2 related factor 2 (NRF2) on ovarian granulosa cells, providing a scientific basis to prevent premature ovarian failure. An ovarian cell injury model was constructed by treating human ovarian granulosa cell (KGN cell) with 4-Vinylcyclohexene dioxide (VCD). Firstly, KGN cells were treated with different concentrations of VCD, and cell counting kit 8 (CCK-8) was used to detect ovarian cell proliferation. After determining IC 50 by CCK8, the levels of estradiol and progesterone in the cell supernatant were detected using enzyme-linked immunosorbent assay (ELISA), reactive oxygen species (ROS) assay kit was used to detect the content of ROS in ovarian cells, real-time fluorescence quantitative polymerase chain reaction (qRT PCR) was used to detect the mRNA expression level of NRF2, and Western blot was used to detect the protein expression level of NRF2. Further, NRF2 silence (siNRF2) and overexpression (NRF2-OE) cell models were constructed through lentivirus transfection, and the effects of regulating NRF2 on VCD treated cell models were investigated by detecting hormone levels, oxidative stress indicators (ROS, SOD, GSH-Px), and autophagy (LC3B level). The results showed that VCD intervention inhibited the proliferation of ovarian granulosa cells in a time-dependent and dose-dependent manner ( F>100, P<0.05), with an IC 50 of 1.2 mmol/L at 24 hours. After VCD treatment, the level of estradiol in the cell supernatant decreased from (56.32±10.18) ng/ml to (24.59±8.75) ng/ml ( t=5.78, P<0.05). Progesterone decreased from (50.25±7.03) ng/ml to (25.13±6.67) ng/ml ( t=6.54, P<0.05). After VCD treatment, the SOD of cells decreased from (44.47±7.71) ng/ml to (30.92±4.97) ng/ml ( t=3.61, P<0.05). GSH-Px decreased from (68.51±10.17) ng/ml to (35.19±6.59) ng/ml ( t=5.73, P<0.05). Simultaneously accompanied by an increase in autophagy and a decrease in NRF2. This study successfully constructed KGN cell models that silenced NRF2 and overexpressed NRF2. Subsequently, this study treated each group of cells with VCD and found that the cell proliferation activity of the siNRF2 group was significantly reduced ( t=8.37, P<0.05), while NRF2-OE could reverse the cell activity damage caused by VCD ( t=3.37, P<0.05). The siNRF2 group had the lowest level of estradiol ( t=5.78, P<0.05), while NRF2-OE could reverse the decrease in cellular estradiol levels caused by VCD ( t=5.58, P<0.05). The siNRF2 group had the lowest progesterone levels ( t=3.02, P<0.05), while NRF2-OE could reverse the decrease in cellular progesterone levels caused by VCD ( t=2.41, P<0.05). The ROS level in the siNRF2 group was the highest ( t=2.86, P<0.05), NRF2-OE could reverse the increase in ROS caused by VCD ( t=3.14, P<0.05), the SOD enzyme content in the siNRF2 group was the lowest ( t=2.98, P<0.05), and NRF2-OE could reverse the decrease in SOD enzyme content caused by VCD ( t=4.72, P<0.05). The GSH-Px enzyme content in the siNRF2 group was the lowest ( t=3.67, P<0.05), and NRF2-OE could reverse the decrease in antioxidant enzyme content caused by VCD ( t=2.71, P<0.05). The LC3B level was highest in the siNRF2 group ( t=2.45, P<0.05), and NRF2-OE was able to reverse the LC3B elevation caused by VCD ( t=9.64, P<0.05). In conclusion, NRF2 inhibits ROS induced autophagy, thereby playing a role in reducing ovarian granulosa cell damage, which may be a potential target for premature ovarian failure.

Objective:To evaluate the predictive value of serum 25-hydroxyvitamin D (25(OH)D) level for the clinical response and imaging response to anal fistula in patients with perianal fistulizing Crohn′s disease (PFCD) treated with ustekinumab (UST).Methods:From October 1, 2021 to June 30, 2023, 80 patients with active PFCD who received UST treatment at the Second Affiliated Hospital of Wenzhou Medical University were retrospectively collected. Harvey-Bradshaw index (HBI) was applied to evaluate the clinical activity of PFCD patients. Perianal disease activity index (PDAI) were used to evaluate the clinical outcomes of anal fistula and pelvic magnetic resonance imaging (MRI) were used to evaluate the imaging outcomes of anal fistula. Serum 25(OH)D levels were examined at week 0, 8, 16, and 24 after UST treatment. Binary logistic regression models were performed to analyze the relationship between the baseline serum 25(OH)D level and the clinical pathological characteristics. And the correlation between the serum 25(OH)D level and the clinical response to anal fistula at week 8 after UST treatment was analyzed. The relationship between clinical response and imaging response to anal fistula at week 24 was also analyzed. R software was employed to draw nomograms and calculate the C-index. Independent sample t test and chi-square test were used for statistical comparison. Results:Multifactorial binary logistic regression analysis showed that the baseline level of serum 25(OH)D was independently correlated with the baseline HBI and baseline PDAI in PFCD patients ( OR=1.45, 95% confidence interval (95% CI) 1.08 to 1.95, P=0.014; OR=1.39, 95% CI 1.01 to 1.92, P=0.042). At week 8 after UST treatment, the serum 25(OH)D level of patients with clinical response to fistula was higher than that of patients without clinical response ((21.77±6.17) μg/L vs. (16.72±6.39) μg/L), while the baseline PDAI was lower than that of patients without response (6.88±2.15 vs. 8.06±2.14), and the proportions of patients with previous failure of biologic therapy and with complex anal fistula were also lower than those of patients without response (42.4%, 14/33 vs. 66.0%, 31/47; 57.6%, 19/33 vs. 78.7%, 37/47), and the differences were statistically significant ( t=3.53 and 2.43, χ2=4.36 and 4.13; P=0.002, 0.022, 0.039 and 0.045). At week 24 after UST treatment, the serum level of 25(OH)D in patients with imaging response was higher than that in patients without response ((22.48±5.81) μg/L vs. (16.66±6.34) μg/L), and the proportion of patients with previous failure of biologic therapy and the proportion of patients with complex anal fistula was lower than that in patients without response (40.0%, 20/50 vs. 12/15; 60.0%, 30/50 vs. 14/15), and all the differences were statistically significant ( t=3.33, χ2=7.39 and 5.86; P=0.004, 0.011 and 0.038). Multifactorial binary logistic regression model analysis showed that the average serum 25(OH)D level and previous failure of biological therapy were 2 independent factors of clinical response to anal fistula at week 8 after UST treatment ( OR=1.11, 95% CI 1.02 to 1.21, P=0.012; OR=0.34, 95% CI 0.12 to 0.97, P=0.043), which were also 2 independent factors of clinical response to anal fistula ( OR=1.14, 95% CI 1.05 to 1.24, P=0.002; OR=0.30, 95% CI 0.11 to 0.89, P=0.029) and imaging response to anal fistula ( OR=1.20, 95% CI 1.05 to 1.36, P=0.006; OR=0.11, 95% CI 0.02 to 0.58, P=0.009) at week 24 after UST treatment. The nomograms showed the C-indexes of the clinical response to anal fistula at week 8 and week 24 after UST treatment were 0.78 (95% CI 0.68 to 0.89) and 0.76 (95% CI 0.64 to 0.87), respectively. The C-index of imaging response at week 24 after UST treatment was 0.85 (95% CI 0.76 to 0.95). Conclusions:In PFCD patients treated with UST, serum 25(OH)D levels and previous failure of biological therapy may independently affect the clinical response to anal fistula at week 8 and 24 after UST treatment, as well as the imaging response to anal fistula at week 24 after UST treatment.

Objective To analyze the level of 25-hydroxyvitamin D[25(OH)D]in patients with type 2 diabetes mellitus,and preliminarily investigate the correlation between serum 25(OH)D level and metabolic indicators such as glycosylated hemoglobin(HbA1c)and pancreatic islet function in patients with type 2 diabetes mellitus.Methods A total of 459 patients with type 2 diabetes mellitus who were hospitalized in the Department of Endocrinology,Xinxiang First People's Hospital from January 2020 to December 2020 were selected as the research subjects.Clinical data of the patients were collected,including gender,age,serum 25(OH)D,fasting insulin,C-peptide,HbA1c,fasting blood glucose,postprandial blood glucose,urinary microalbumin,urinary albumin-to-creatinine ratio,blood calcium,blood uric acid(UA),triglyceride(TG),total cholesterol(TCH),low-density lipoprotein(LDL),and high-density lipoprotein(HDL).According to the serum 25(OH)D level,the patients were divided into sufficient group[n=20,25(OH)D≥30 μg·L-1],insufficient group[n=95,20 μg·L-125(OH)D＜30 μg·L-1],deficient group[n=231,10 μg·L-1 ≤25(OH)D＜20 μg·L-1],and severely deficient group[n=113,25(OH)D＜10 μg·L-1].Differences in metabolic indicators of patients in the four groups were compared,and the correlation between 25(OH)D level and metabolic indicators was analyzed by using the Pearson correlation.Results The serum 25(OH)D level of patients with type 2 diabetes mellitus was 3.00-46.59(15.75±0.35)μg·L-1;the serum 25(OH)D level of male patients was significantly higher than that of female patients(P＜0.05).The prevalence of 25(OH)D deficiency in patients with type 2 diabetes mellitus was 74.9％(344/459).The 25(OH)D deficiency mainly occurred in January,February,March,April,November,and December.Patients in the insufficient,deficient,and severely deficient groups had significantly higher HbA1c levels than those in the sufficient group(P＜0.05),and the HbA1c levels of patients in the deficient and severely deficient groups were significantly higher than those in the insufficient group(P＜0.05);there was no statistically significant difference in the HbA1c level between the deficient group and the severely deficient group(P＞0.05).There was no statistically significant difference in fasting blood glucose between the sufficient group and insufficient group,and between the deficient group and severely deficient group(P＞0.05);fasting blood glucose of patients in the deficient and severely deficient groups was significantly higher than that in the sufficient and insufficient groups(P＜0.05).There was no statistically significant difference in fasting insulin,urinary microalbumin,daily total urinary albumin,and urinary albumin-to-creatinine ratio of patients among the sufficient,insufficient,and deficient groups(P＞0.05);the fasting insulin of patients in the severely deficient group was significantly lower than that in the sufficient,insufficient,and deficient groups(P＜0.05);the urinary microalbumin,daily total urinary albumin,and urinary albumin-to-creatinine ratio of patients in the severely deficient group were significantly higher than those in the sufficient,insufficient,and deficient groups(P＜0.05).There was no statistically significant difference in the homeostasis model assessment for insulin resistance(HOMA-IR),serum albumin,blood creatinine,1-hour postprandial blood glucose,2-hour postprandial blood glucose,3-hour postprandial blood glucose,fasting C-peptide,1-hour postprandial C-peptide,2-hour postprandial C-peptide,3-hour postprandial C-peptide,TG,TCH,LDL,HDL,blood UA,and blood calcium of patients among the four groups(P＞0.05).Pearson correlation analysis showed that serum 25(OH)D levels in patients with type 2 diabetes mellitus were negatively correlated with HbA1c,urinary microalbumin,and urinary albumin-to-creatinine ratio(r=-0.093,-0.166,-0.157;P＜0.05),and positively correlated with fasting insulin(r=0.089,P＜0.05).Serum 25(OH)D levels in patients with type 2 diabetes mellitus had no correlation with fasting blood glucose,HOMA-IR,serum albumin,blood creatinine,1-hour postprandial blood glucose,2-hour postprandial blood glucose,3-hour postprandial blood glucose,fasting C-peptide,1-hour postprandial C-peptide,2-hour postprandial C-peptide,3-hour postprandial C-peptide,TG,TCH,LDL,HDL,blood UA,and blood calcium(P＞0.05).Conclusion 25(OH)D deficiency and insufficiency are common in patients with type 2 diabetes mellitus,especially in female patients.In patients with type 2 diabetes mellitus,25(OH)D level is positively correlated with fasting insulin and negatively correlated with HbAlc,urinary microalbumin,and urinary albumin-to-creatinine ratio.25(OH)D deficiency in patients with type 2 diabetes mellitus is mainly distributed in January,February,March,April,November,and December.

This study explores the effects and possible mechanisms of nuclear factor E2 related factor 2 (NRF2) on ovarian granulosa cells, providing a scientific basis to prevent premature ovarian failure. An ovarian cell injury model was constructed by treating human ovarian granulosa cell (KGN cell) with 4-Vinylcyclohexene dioxide (VCD). Firstly, KGN cells were treated with different concentrations of VCD, and cell counting kit 8 (CCK-8) was used to detect ovarian cell proliferation. After determining IC 50 by CCK8, the levels of estradiol and progesterone in the cell supernatant were detected using enzyme-linked immunosorbent assay (ELISA), reactive oxygen species (ROS) assay kit was used to detect the content of ROS in ovarian cells, real-time fluorescence quantitative polymerase chain reaction (qRT PCR) was used to detect the mRNA expression level of NRF2, and Western blot was used to detect the protein expression level of NRF2. Further, NRF2 silence (siNRF2) and overexpression (NRF2-OE) cell models were constructed through lentivirus transfection, and the effects of regulating NRF2 on VCD treated cell models were investigated by detecting hormone levels, oxidative stress indicators (ROS, SOD, GSH-Px), and autophagy (LC3B level). The results showed that VCD intervention inhibited the proliferation of ovarian granulosa cells in a time-dependent and dose-dependent manner ( F>100, P<0.05), with an IC 50 of 1.2 mmol/L at 24 hours. After VCD treatment, the level of estradiol in the cell supernatant decreased from (56.32±10.18) ng/ml to (24.59±8.75) ng/ml ( t=5.78, P<0.05). Progesterone decreased from (50.25±7.03) ng/ml to (25.13±6.67) ng/ml ( t=6.54, P<0.05). After VCD treatment, the SOD of cells decreased from (44.47±7.71) ng/ml to (30.92±4.97) ng/ml ( t=3.61, P<0.05). GSH-Px decreased from (68.51±10.17) ng/ml to (35.19±6.59) ng/ml ( t=5.73, P<0.05). Simultaneously accompanied by an increase in autophagy and a decrease in NRF2. This study successfully constructed KGN cell models that silenced NRF2 and overexpressed NRF2. Subsequently, this study treated each group of cells with VCD and found that the cell proliferation activity of the siNRF2 group was significantly reduced ( t=8.37, P<0.05), while NRF2-OE could reverse the cell activity damage caused by VCD ( t=3.37, P<0.05). The siNRF2 group had the lowest level of estradiol ( t=5.78, P<0.05), while NRF2-OE could reverse the decrease in cellular estradiol levels caused by VCD ( t=5.58, P<0.05). The siNRF2 group had the lowest progesterone levels ( t=3.02, P<0.05), while NRF2-OE could reverse the decrease in cellular progesterone levels caused by VCD ( t=2.41, P<0.05). The ROS level in the siNRF2 group was the highest ( t=2.86, P<0.05), NRF2-OE could reverse the increase in ROS caused by VCD ( t=3.14, P<0.05), the SOD enzyme content in the siNRF2 group was the lowest ( t=2.98, P<0.05), and NRF2-OE could reverse the decrease in SOD enzyme content caused by VCD ( t=4.72, P<0.05). The GSH-Px enzyme content in the siNRF2 group was the lowest ( t=3.67, P<0.05), and NRF2-OE could reverse the decrease in antioxidant enzyme content caused by VCD ( t=2.71, P<0.05). The LC3B level was highest in the siNRF2 group ( t=2.45, P<0.05), and NRF2-OE was able to reverse the LC3B elevation caused by VCD ( t=9.64, P<0.05). In conclusion, NRF2 inhibits ROS induced autophagy, thereby playing a role in reducing ovarian granulosa cell damage, which may be a potential target for premature ovarian failure.

ObjectiveTo establish a rapid and stable liquid chromatography-mass spectrometry（LC-MS） for simultaneous analysis of 17 chemical components in Gnaphalium affine aboveground parts with flowers， so as to provide experimental basis for improving the quality standard of this herb. MethodUltra performance liquid chromatography-quadrupole/electrostatic field orbitrap mass spectrometry（UPLC-Q-Exactive Orbitrap MS） was used for the quantitative analysis of 17 constituents in 15 batches of G. affine from different origins， the separation was performed on an ACQUITY UPLC^® BEH C₁₈ column（2.1 mm×100 mm， 1.7 μm） with the mobile phase of methanol（A）-0.1% formic acid aqueous solution（B） for gradient elution（0-1.0 min， 8%A； 1.0-4.0 min， 8%-26%A； 4.0-9.0 min， 26%A； 9.0-14.0 min， 26%-34%A； 14.0-14.5 min， 34%-45%A； 14.5-15.0 min， 45%-60%A； 15.0-18.0 min， 60%-90%A； 18.0-19.0 min， 90%A； 19.0-19.01 min， 90%-8%A； 19.01-20.0 min， 8%A）， the flow rate was 0.3 mL·min^-1， the column temperature was 40 ℃ and the injection volume was 2 μL. And the electrospray ionization was used with full scanning in both positive and negative ion modes， and the scanning range was m/z 100-1 000. ResultThe established method has been verified by the methodology and could be used for the simultaneous quantification of 17 components in G. affine. The content ranges of the 17 components（quinic acid， gallic acid， protocatechuic acid， neochlorogenic acid， chlorogenic acid， cryptochlorogenic acid， caffeic acid， 1，3-O-dicaffeoylquinic acid， isochlorogenic acid A， isoquercitrin， 1，5-O-dicaffeoylquinic acid， apigenin-7-O-glucoside， astragalin， isochlorogenic acid C， luteolin， apigenin and hispidulin） in 15 batches of G. affine samples was 39.60-179.12， 0.17-0.84， 2.41-8.38， 4.33-31.50， 13.63-180.38， 2.43-14.75， 1.16-19.68， 0.49-5.63， 55.77-445.16， 0.23-10.26， 62.04-530.10， 1.11-18.01， 11.36-90.61， 12.22-65.98， 7.22-69.84， 3.37-45.65， 0.30-2.59 μg·g^-1， respectively. The content of organic acids was higher than that of flavonoids in G. affine， and the contents of 1，5-O-dicaffeoylquinic acid， isochlorogenic acid A， quinic acid and chlorogenic acid were higher. Meanwhile， the content of flavonoids in the samples from Guizhou was higher than that from Jiangsu， while the content of organic acids in the samples from Jiangsu was higher than that from Guizhou. ConclusionThe established method can be used for the rapid and accurate determination of 17 components in G. affine， which clarifies the content range of the main components in this herb， and can provide a reference for the selection of quality control markers of G. affine.

Currently,the research or publications related to the clinical comprehensive evaluation of Chinese patent medicine are increasing,which attracts the broad attention of all circles. According to the completed clinical evaluation report on Chinese patent medicine,there are still practical problems and technical difficulties such as unclear responsibility of the evaluation organization,unclear evaluation subject,miscellaneous evaluation objects,and incomplete and nonstandard evaluation process. In terms of evaluation standards and specifications,there are different types of specifications or guidelines with different emphases issued by different academic groups or relevant institutions. The professional guideline is required to guide the standardized and efficient clinical comprehensive evaluation of Chinese patent medicine and further improve the authority and quality of evaluation. In combination with the characteristics of Chinese patent medicine and the latest research achievement at home and abroad,the detailed specifications were formulated from six aspects including design,theme selection,content and index,outcome,application and appraisal,and quality control. The guideline was developed based on the guideline development requirements of China Assoication of Chinese medicine. After several rounds of expert consensus and public consultation,the current version of the guideline has been developed.
Medicine, Chinese Traditional ; Nonprescription Drugs ; Consensus ; China ; Reference Standards ; Drugs, Chinese Herbal

OBJECTIVE: To explore and verify that transient receptor potential vanilloid 4(TRPV4) affects chondrocyte degeneration. METHODS: Neonatal SD rats were selected, primary chondrocytes were extracted, and identified by toluidine blue staining and alcian blue staining;an in vitro chondrocyte inflammation model was constructed by IL-1β, and TRPV4 inhibitor was used to treat chondrocytes under inflammatory conditions, and the chondrocytes were treated by RT-PCR method was used to detect matrix metallopeptidase 13(MMP-13), a disintegrin and metalloproteinase with thrombospondin 5, (ADAMTS-5)、nitric oxide synthase 2(NOS2)、Collagen, type II alpha 1(Col2α1)and aggrecan (Acan) mRNA in chondrocytes; primary chondrocytes were treated with different concentrations of TRPV4 overexpression plasmid, and the optimal overexpression dose was screened. The mRNA expressions of TRPV4, MMP-13, ADAMTS-5, NOS2, Col2α1 and Acan in chondrocytes under the optimal TRPV4 overexpression dose were detected. RESULTS: Toluidine blue staining and Alcian blue staining identified the extracted cells as primary chondrocytes;RT-PCR showed that TRPV4, MMP-13, ADAMTS-5, NOS2 mRNA in chondrocytes treated with TRPV4 inhibitor under inflammatory conditions. The expression of Col2α1 mRNA was significantly decreased (P<0.05), and the expression of Col2α1 mRNA was increased (P<0.05). Although there was no significant difference in the expression of Acan mRNA, the overall trend was also increasing. The expression of Col2α1 and Acan mRNA in chondrocytes was significantly decreased (P<0.05), and the expression of NOS2 mRNA was increased(P<0.05), but there was no significant difference in MMP-13 and ADAMTS-5 (P>0.05). CONCLUSION Inhibiting the expression of TRPV4 can down-regulate the expression of genes related to chondrocyte degeneration.
Animals ; Rats ; Aggrecans/metabolism* ; Cartilage, Articular ; Cells, Cultured ; Chondrocytes ; Interleukin-1beta/metabolism* ; Matrix Metalloproteinase 13/metabolism* ; Rats, Sprague-Dawley ; RNA, Messenger/metabolism* ; TRPV Cation Channels/metabolism*