ObjectiveThis study aimed to evaluate the clinical efficacy of Ruyi Zhenbaowan（RYZBW）in the treatment of initial and early knee osteoarthritis （KOA） through a prospective multicenter，randomized，double-blind，and placebo-parallel controlled trial. MethodFrom October 13^th， 2021 to December 25^th， 2021， 240 KOA subjects meeting the acceptance criteria were enrolled in 15 sub-centers including Wangjing Hospital， Chinese Academy of Chinese Medical Sciences， and they were randomly divided into observation group and control group， with 120 cases in each group. The intervention measures for the observation group were RYZBW + health education， and the intervention measures for the control group were RYZBW placebo + health education. The intervention period in both groups was four weeks， and they were followed up for four weeks after the intervention. The primary outcome measure was the total score of Western Ontario and McMaster University Osteoarthritis Index score （WOMAC score）， and the secondary outcome measures were the response rate of visual scale （VAS） pain score， WOMAC sub item scores （joint pain， joint stiffness， and joint function）， quality of life （SF-12） score， and traditional Chinese medicine （TCM） syndrome score. Result（1） Efficacy evaluation. The marginal model results showed that the observation group was better than the control group in improving the WOMAC total score and WOMAC pain score in the treatment of KOA with RYZBW， and the difference was statistically significant （P<0.05）. There was no significant difference between the two groups in improving VAS score response rate， WOMAC function score， WOMAC stiffness score， SF12-PCS （quality of life-physical health） score， SF12-MCS （quality of life-mental health） score， and TCM syndrome score. （2） Subgroup analysis. ① In terms of VAS score response rate， the response rate of the observation group was higher than that of the control group for subjects with baseline VAS score of （4， 5］， and the difference was statistically significant （P<0.05）. ② In terms of TCM syndrome score， for subjects aged ［56， 60］ and ［61， 65］， the decrease in total TCM syndrome score in the observation group was better than that in the control group， and the difference was statistically significant （P<0.05）. ConclusionTibetan medicine RYZBW has good clinical efficacy in improving WOMAC total score， VAS score response rate， WOMAC pain score， WOMAC function score， and TCM syndrome score for patients with initial and early KOA， which can fill the lack of Tibetan medicine RYZBW in the treatment of KOA and make a demonstration study for the inheritance and development of ethnic medicine.

Objective To investigate the expression of miR-6768-5p in lung cancer tissue and its effect on the proliferation and invasion of lung cancer cells through targeted regulation of carboxypeptidase A4(CPA4).Methods The expression of miR-6768-5p in lung cancer tissues and adjacent tissues was analyzed u-sing the TCGA database.Quantitative real-time PCR(qPCR)was used to detect the expression of miR-6768-5p in human lung cancer cell lines(HCC1588,H1650,H1299,A549,HCC827)and normal alveolar epithelial cells(HPAEpiC cells).Lung cancer cells were transfected with NC mimics and miR-6768-5p mimics,respec-tively,and divided into NC group and miR-6768-5p group.The MTS assay and Matrigel invasion assay were used to detect the cell proliferation and invasion ability of each group,respectively.The putative binding sites of miR-6768-5p and CPA4 were verified using RNAhybrid software and dual-luciferase reporter gene experi-ment.The expression of CPA4 mRNA in each group of cells was detected by qPCR.The expression of AKT/c-MYC signaling pathway proteins in the cells of each group was analyzed by Western blot.Results Com-pared with the adjacent tissues,the relative expression level of miR-6768-5p in lung cancer tissues was signifi-cantly decreased,and the difference was statistically significant(P＜0.05).Compared with HPAEpiC cells,the relative expression level of miR-6768-5p was significantly decreased in lung cancer cell lines,and the differ-ence was statistically significant(P＜0.05).Compared with the NC group,the cell proliferation rate of miR-6768-5p group was significantly decreased(P＜0.05).The number of invasive cells in NC group and miR-6768-5p group was(131.30±12.55)and(37.45±7.77),respectively,and the number of invasive cells in miR-6768-5p group was significantly lower than that in NC group(P＜0.05).The relative expression level of CPA4 mRNA in H1299 cells of miR-6768-5p group was significantly lower than that in NC group(t=4.93,P＜0.05).Compared with the NC group,the expressions of AKT/c-myC signaling pathway proteins p-AKT,p-mTORC1,XIAP,MDM2 and C-myC proteins in miR-6768-5p group were significantly decreased.Conclusion The expression of miR-6768-5p is decreased in lung cancer tissues,and miR-6768-5p may inhibit the activation of AKT/c-MYC signaling pathway by targeting CPA4,and reduce the proliferation and invasion ability of lung cancer H1299 cells.

Objective:To analyze the research status of scleral lenses by bibliometric method based on the relevant collection of scleral lenses in the Web of Science database.Methods:Using the scleral lens-related literature collected in the Web of Science Core Collection database in the past 10 years (January 2013 to December 2022) as the object of analysis, bibliometric method and CiteSpace tools were used to conduct visual analysis of the literature.A comprehensive analysis of the volume of literature published, the distribution of countries and institutions, the information of core authors, the distribution of journals, and keyword clustering was performed.Results:A total of 340 articles were retrieved, which were published in 54 journals, with an average of 6.3 articles per journal, involving 301 authors.Research in this area covered 35 countries or regions, and 256 research institutions were involved.Discipline development was mainly in the United States, India, Spain and Australia.The main focus was on scleral lens (scleral contact lens), ocular surface disease, corneal edema, miniature scleral lens, etc.In the past 10 years, the trend of research hot topics in scleral lenses had shifted from the initial study of combining scleral contact lenses with ocular surface diseases to the subsequent study of prosthetic replacement of the ocular surface ecosystem, and the exploration of corneal clearance and shape.From 2013 to 2021, the main focuses were ocular surface diseases, scleral contact lenses, and corneal edema.After 2021, research on ocular surface diseases and keratoplasty declined.From January 2013 to December 2022, emergent keywords related to scleral lens mainly included scleral contact lens, transplantation, anti-host disease, prosthetic device in the first stage, artificial replacement of ocular surface ecosystem and irregular cornea in the second stage, and the research on corneal gap and characteristic shape in the third stage.Optical coherence tomography and corneal topography were commonly used examinations for scleral lens research and fitting.Conclusions:At present, the scleral lens is mainly used for dry eye, corneal diseases, corneal ectasia, keratitis, and corneal transplantation, especially after penetrating keratoplasty and refractive errors.Prosthetic replacement of the ocular surface ecosystem, and the exploration of corneal clearance and shape are the research hotspots in scleral lenses.

Objective:To investigate the correlation between fat distribution and the composite indices of femoral neck strength in obese postmenopausal women.Methods:A total of 293 postmenopausal women with non-low body weight were selected, laboratory tests, body composition analyzer test and double-energy X-ray absorptiometry scan were performed. Based on the body mass index(BMI), they were divided into three groups, the normal BMI group(18.5 kg/m 2≤BMI<24.0 kg/m 2, n=91), the overweight group(24.0 kg/m 2≤BMI<28.0 kg/m 2, n=115), and the obese group(BMI≥28.0 kg/m 2, n=87). The measurement results were analyzed. Results:In the obese group, bone mineral density(BMD) of all sites was higher than that in the normal BMI group and overweight group( P<0.005), compression strength index(CSI), bending strength index(BSI), and impact strength index(ISI) were significantly lower than those in the normal BMI group( P<0.001, P=0.008, P=0.001). In the obese group, waist circumference, waist-hip ratio, total fat mass, appendicular fat mass, and trunk fat mass were risk factors for CSI, BSI and ISI independent of age, fasting blood glucose, and BMI( P<0.05). Visceral fat grade and Chinese visceral adiposity fat index were the risk factors for CSI, BSI, and ISI( P<0.05). Conclusion:The composite indices of femoral neck strength decreased in obese postmenopausal women, and both subcutaneous fat and visceral fat were negatively associated with the composite indices of femoral neck strength.

Background Permethrin is a commonly used pyrethroid insecticide and has been found to be potentially neurotoxic. Microglia are innate immune cells in the central nervous system and are involved in the development of a range of neurodegenerative diseases. Objective To observe possible toxic effects of permethrin on human microglia clone 3 (HMC3) in vitro and explore associated mechanism. Methods HMC3 were treated with 0, 10, 25, and 55 μmol·L⁻¹ permethrin for 72 h. Cell cycle and apoptosis were measured using flow cytometry. Cyclin-dependent kinase 1 (CDK1), cyclin-dependent kinase inhibitor 1A (CDKN1A), cyclin B2 (CCNB2), cellular tumor antigen p53 (p53), factor-related apoptosis (FAS), caspase 3 (CASP3), and H2A histone family member X (H2AX) were detected by quantitative real-time PCR (qPCR). The differential genes and enrichment pathways of HMC3 after 0 and 25 μmol·L⁻¹ permethrin treatment was analyzed by RNA sequencing. HMC3 was treated by 0, 10, 25, and 55 μmol· L⁻¹ permethrin for 72 h. The content of nitric oxide (NO) in the supernatant was detected using Griess reagent. The secretion level of interleukin-6 (IL-6) was detected by enzyme linked immunosorbent assay (ELISA). The mRNA expression levels of mitogen-activated protein kinase (MAPK) pathway (including MAPK1, MAPK8, and MAPK14), interleukin-1β (IL-1β), IL-6, and matrix metalloproteinase (MMP) families (including MMP1, MMP2, MMP3, and MMP9) were detected by qPCR. The protein expressions of phosphorylated p38 mitogen-activated protein kinase (p-p38), phosphorylated extracellular signal-regulated kinase (p-ERK), IL-1β, IL-6, and MMP1 were detected by Western blot. Results HMC3 was arrested in G2/M phase after 0, 10, 25, and 55 μmol·L⁻¹ permethrin treatment for 72 h, of which there was a statistically significant difference between the 55 μmol·L⁻¹ permethrin treatment group and the control group (P<0.01), and the mRNA expression of CDKN1A was up-regulated according to the qPCR (P<0.05). There was no statistically significant difference in the proportions of apoptosis between the groups (P＞0.05). The RNA sequencing showed that the differential genes were enriched in the MAPK pathway, and the mRNA expressions of MAPK1, MAPK8, and MAPK14 were up-regulated after the permethrin treatment at 55 μmol·L⁻¹ compared to the control group by qPCR (P<0.05). The Western blot revealed that, compared to the control group, the levels of p-p38 and p-ERK were increased after the 10 μmol·L⁻¹ permetrin treatment (P<0.05), the p-ERK level was increased after the 25 μmol·L⁻¹ permetrin treatment (P<0.05), and the p-p38 level was up-regulated after the 55 μmol·L⁻¹ permetrin treatment (P<0.05). The secretion of NO in the supernatant of HMC3 increased after permetrin treatment compared to the control group (P<0.05), the mRNA and protein expressions and the secretion of IL-6 showed an upward trend, the mRNA and protein expressions of IL-1β were up-regulated (P<0.05), and the mRNA and protein expressions of MMP1 were up-regulated in the 25 and 55 μmol·L⁻¹ permethrin groups (P<0.05). Conclusion Permethrin inhibits HMC3 cell proliferation in vitro, induces cell cycle arrest, activates MAPK pathway, and promotes the expression of inflammatory factors IL-1β and MMP1, which may be one of the mechanism of neurotoxicity induced by permethrin.

Flavanone 3-hydroxylase (F3H) is a key enzyme in the synthesis of phycocyanidins. In this experiment, the petals of red Rhododendron hybridum Hort. at different developmental stages were used as experimental materials. The R. hybridum flavanone 3-hydroxylase (RhF3H) gene was cloned using reverse transcription PCR (RT-PCR) and rapid-amplification of cDNA ends (RACE) techniques, and bioinformatics analyses were performed. Petal RhF3H gene expression at different developmental stages were analyzed by using quantitative real-time polymerase chain reaction (qRT-PCR). A pET-28a-RhF3H prokaryotic expression vector was constructed for the preparation and purification of RhF3H protein. A pCAMBIA1302-RhF3H overexpression vector was constructed for genetic transformation in Arabidopsis thaliana by Agrobacterium-mediated method. The results showed that the R. hybridum Hort. RhF3H gene is 1 245 bp long, with an open reading frame of 1 092 bp, encoding 363 amino acids. It contains a Fe²⁺ binding motif and a 2-ketoglutarate binding motif of the dioxygenase superfamily. Phylogenetic analysis showed that the R. hybridum RhF3H protein is most closely related to the Vaccinium corymbosum F3H protein. qRT-PCR analysis showed that the expression level of the red R. hybridum RhF3H gene tended to increase and then decrease in the petals at different developmental stages, with the highest expression at middle opening stage. The results of the prokaryotic expression showed that the size of the induced protein of the constructed prokaryotic expression vector pET-28a-RhF3H was about 40 kDa, which was similar to the theoretical value. Transgenic RhF3H Arabidopsis thaliana plants were successfully obtained, and PCR identification and β-glucuronidase (GUS) staining demonstrated that the RhF3H gene was integrated into the genome of A. thaliana plants. qRT-PCR, total flavonoid and anthocyanin contentanalysis showed that RhF3H was significantly higher expressed in the transgenic A. thaliana relative to that of the wild type, and its total flavonoid and anthocyanin content were significantly increased. This study provides a theoretical basis for investigating the function of RhF3H gene, as well as for studying the molecular mechanism of flower color in R. simsiib Planch.
Arabidopsis/metabolism* ; Rhododendron/metabolism* ; Amino Acid Sequence ; Anthocyanins/metabolism* ; Phylogeny ; Flavonoids/metabolism* ; Cloning, Molecular ; Gene Expression Regulation, Plant ; Plant Proteins/metabolism*

Objective:To investigate the clinical features, diagnosis, treatment, and future sex selection in patients with partial androgen insensitivity syndrome (PAIS).Methods:Retrospective analysis of clinical data of three patients with PAIS who received treatment in The Third Affiliated Hospital of Xi'an Medical University, Beijing Ditan Hospital of Capital Medical University, and Beijing Yayuncun Amcare Women's and Children's Hospital from 2013 to 2015 was conducted. Physical signs, specialized examinations, surgical explorations, and treatments were analyzed. The Chinese database was searched, and 12 cases of PAIS were collected and summarized.Results:Fifteen patients with PAIS presented with primary amenorrhea (15/15). Special clinical manifestations included gender as male or appearing as male (5/15), penile dysplasia or clitoral hypertrophy (14/15), urethral dysplasia (5/15), and breast development (4/15). Eleven cases were treated based on female gender (including surgery and hormone replacement therapy). There were three special patients with PAIS who had specific etiology, genetics, clinical manifestations, histopathology, diagnosis, and treatment and ultimately underwent treatment based on female gender.Conclusion:PAIS is a rare form of disorder of sex development, featuring a karyotype of 46, XY, and is a congenital X-linked recessive condition. Understanding the pathogenesis of PAIS more thoroughly can contribute to accurate diagnosis, personalized treatment, and well-organized follow-up, thereby preventing gender dysphoria.

Objective:To assess the combination of conventional ultrasound with contrast-enhanced ultrasound(CEUS)in the diagnosis of urothelial carcinoma(UC)of the renal pelvis in elderly patients.Methods:Sixty-seven elderly patients with a histopathologic diagnosis of UC of the renal pelvis and surgically treated at our hospital between April 2015 and March 2023 were retrospectively analyzed.Characteristics of regular preoperative 2D ultrasound, color Doppler flow imaging(CDFI)and CEUS were examined.Results:Of 67 patients, 49(73.13%)were found to have localized lesions in the renal pelvis and renal calyces.Lesions in 53 patients(79.10%)could be clearly identified by conventional ultrasound, with 46(86.79%)being isoechoic or hypoechoic, and 7(13.21%)being hyperechoic.Analysis of tumor blood flow by CDFI found 22 cases(41.51%)with avascular lesions, 21(39.62%)with hypovascular lesions and 10(18.87%)with hypervascular lesions.The average value of the resistance index(RI)was 0.64.Enhancement was seen in 62 lesions(92.54%)by CEUS after injection of SonoVue.Compared with the cortex of the ipsilateral kidney, a slow enhancement pattern was observed in 46(74.19%), 14(22.58%)showed simultaneous enhancement, and 2(3.23%)showed fast enhancement.At peak enhancement, 43 lesions(69.35%)had hypo-enhancement, 10(16.13%)had iso-enhancement, and 9(14.52%)had hyper-enhancement, compared with the cortex.Concerning the homogeneity of enhancement, 16(25.81%)displayed heterogeneous enhancement, with tumor necrosis or hemorrhage, and 46(74.19%)had homogeneous enhancement.When the contrast agent washout rate was assessed, a fast washout pattern was observed in 53(85.48%), synchronous washout in 6(9.68%), and slow washout in 3(4.84%).Conclusions:UC of the renal pelvis mostly shows isoechoic and hypoechoic lesions on conventional ultrasound, avascular or hypo-vascular lesions on CDFI, and slow-in, fast-out and hypo-enhancement on CEUS, compared with the cortex.Conventional ultrasound combined with CEUS can help improve the diagnostic accuracy of UC of the renal pelvis.

Objective:To evaluate the effect of ozone preconditioning on splenic natural killer (NK) cells in septic mice.Methods:Twenty-four SPF-grade male C57BL/6 mice, aged 6-8 weeks, weighing 18-22 g, were divided into 4 groups ( n=6 each) according to the random number table method: control group (group C), lipopolysaccharide (LPS)group, ozone+ LPS group (O 3+ LPS) and air+ LPS group (Air+ LPS). The sepsis was induced by intraperitoneal injection of LPS 10 mg/kg. Ozone preconditioning was started at 5 days before developing the model: ozone 1 mg/kg was intraperitoneally injected once a day for 5 consecutive days, the equal volume of air was injected in Air+ LPS group. The survival was observed within 72 h after LPS injection, and sepsis score and ear temperature (once every 2 h, an average was calculated) were recorded. The posterior orbital venous blood samples were taken at 6 and 24 h after LPS injection for determination of serum interferon-γ (IFN-γ) and interleukin-10 (IL-10) concentrations using enzyme-linked immunosorbent assay. The spleen was then taken, and a single cell suspension of the spleen was prepared for measurement of the percentage of NK cells in the spleen by flow cytometry. Results:Compared with C group, the ear temperature, sepsis score and 72-h survival rate were significantly decreased, serum IFN-γ and IL-10 concentrations were increased at each time point after LPS injection, and the percentage of splenic NK cells was increased at 6 h after LPS injection and decreased at 24 h after LPS injection in LPS, Air+ LPS and O 3+ LPS groups ( P<0.05). Compared with LPS group, the ear temperature, sepsis score and 72-h survival rate were significantly increased, serum IFN-γ concentrations were decreased at each time point after LPS injection, serum IL-10 concentrations were increased at each time point after LPS injection, and the percentage of splenic NK cells was decreased at 6 h after LPS injection and increased at 24 h after LPS injection in O 3+ LPS group ( P<0.05), and no significant change was found in the parameters mentioned above in Air+ LPS group ( P>0.05). Conclusions:The mechanism by which ozone preconditioning reduces sepsis may be related to reduction of inflammatory responses and regulation of splenic NK cell levels in septic mice.

Seed quality plays an important role in the agricultural and animal husbandry production, the effective utilization of genetic resources, the conservation of biodiversity and the restoration and reconstruction of plant communities. Seed aging is a common physiological phenomenon during storage. It is a natural irreversible process that occurs and develops along with the extension of seed storage time. It is not only related to the growth, yield and quality of seed and seedling establishment, but also has an important effect on the conservation, utilization and development of plant germplasm resources. The physiological mechanisms of seed aging are complex and diverse. Most studies focus on conventional physiological characterization, while systematic and comprehensive in-depth studies are lacking. Here we review the recent advances in understanding the physiology of seed aging process, including the methods of seed aging, the effect of aging on seed germination, and the physiological and molecular mechanisms of seed aging. The change of multiple physiological parameters, including seed vigor, electrical conductivity, malondialdehyde content and storage material in the seed, antioxidant enzyme activity and mitochondrial structure, were summarized. Moreover, insights into the mechanism of seed aging from the aspects of transcriptome, proteome and aging related gene function were summarized. This study may facilitate the research of seed biology and the conservation and utilization of germplasm resources.
Germination ; Plants ; Proteome ; Seedlings ; Seeds/genetics*