G protein-coupled receptor kinase 2 (GRK2) participates in the phosphorylation and desensitization of G protein-coupled receptor (GPCR), impacting various biological processes such as inflammation and cell proliferation. Dysregulated expression and activity of GRK2 have been reported in multiple cells in rheumatoid arthritis (RA). However, whether and how GRK2 regulates synovial hyperplasia and fibroblast-like synoviocytes (FLSs) proliferation is poorly understood. In this study, we investigated the regulation of GRK2 and its biological function in RA. We found that GRK2 transmembrane activity was increased in FLSs of RA patients and collagen-induced arthritis (CIA) rats. Additionally, we noted a positive correlation between high GRK2 expression on the cell membrane and serological markers associated with RA and CIA. Immunoprecipitation-mass spectrometry and pull-down analyses revealed tumor necrosis factor receptor-associated factor 2 (TRAF2) as a novel substrate of GRK2. Furthermore, surface plasmon resonance (SPR) and molecular docking assays determined that the C-terminus of GRK2 binds to the C-terminus of TRAF2 at the Gln340 residue. GRK2 knockdown and the GRK2 inhibitor CP-25 attenuated synovial hyperplasia and FLS proliferation in CIA both in vitro and in vivo by decreasing GRK2 membrane expression and activity. Mechanistically, increased GRK2 transmembrane activity contributed to the recruitment of TRAF2 on the cell membrane, promoting GRK2-TRAF2 interactions that facilitate the recruitment of the E3 ubiquitin ligase TRIM47 to TRAF2. This enhanced TRAF2 Lys63 polyubiquitylation and induced nuclear factor (NF)-κB activation, leading to synovial hyperplasia and abnormal proliferation of FLSs. Our study provides a mechanistic and preclinical rationale for further evaluation of GRK2 as a therapeutic target for RA.

The rapid development of artificial intelligence (AI), especially generative AI (GenAI), has already brought, and will continue to bring, revolutionary changes to our daily production and life, as well as create new opportunities and challenges for diagnostic and therapeutic practices in the medical field. Haihe Hospital of Tianjin University collaborates with the National Supercomputer Center in Tianjin, Tianjin University, and other institutions to carry out research in areas such as smart healthcare, smart services, and smart management. We have conducted research and development of a full-process disease diagnosis and treatment assistance system based on GenAI in the field of smart healthcare. The development of this project is of great significance. The first goal is to upgrade and transform the hospital's information center, organically integrate it with existing information systems, and provide the necessary computing power storage support for intelligent services within the hospital. We have implemented the localized deployment of three models: Tianhe "Tianyuan", WiNGPT, and DeepSeek. The second is to create a digital avatar of the chief physician/chief physician's voice and image by integrating multimodal intelligent interaction technology. With generative intelligence as the core, this solution provides patients with a visual medical interaction solution. The third is to achieve deep adaptation between generative intelligence and the entire process of patient medical treatment. In this project, we have developed assistant tools such as intelligent inquiry, intelligent diagnosis and recognition, intelligent treatment plan generation, and intelligent assisted medical record generation to improve the safety, quality, and efficiency of the diagnosis and treatment process. This study introduces the content of a full-process disease diagnosis and treatment assistance system, aiming to provide references and insights for the digital transformation of the healthcare industry.
Artificial Intelligence ; Humans ; Delivery of Health Care ; Generative Artificial Intelligence

Objective To construct a stable synovial cell line MH7A from rheumatoid arthritis(RA)patients using lentiviral vectors that interfere with the expression of tumor necrosis factor receptor associated factor 2(TRAF2),and to study the role of TNF-α-TRAF2 signaling in MH7A abnormal proliferation.Methods Based on the design principles of human TRAF2 gene sequence and shRNA sequence,three pairs of TRAF2 shRNA interference se-quences were designed and synthesized.The primers were annealed by PCR,and a linear vector was obtained by double enzyme digestion PLKO.1-puro.The linearized vector was connected to the annealed primers through Solu-tion I,and the connected products were introduced into receptive cells.The plates were coated,and positive colo-nies were selected for sequencing.Three different recombinant plasmids of PLKO.1-TRAF2-shRNA lentivirus were constructed,and lentivirus packaging plasmids was used to package logarithmic growth phase HEK 293T cells.Vi-rus solution was collected to infect MH7A cells.At the same time,puromycin was used to screen MH7A stable transgenic strains with low TRAF2 expression.CCK-8 method,Western blot,and qPCR were used to detect the proliferation function of MH7A induced by TNF-α and low expression of TRAF2,as well as downstream signal TRAF2,P65 protein expression and mRNA levels.Results PLKO.1-TRAF2-shRNA(1),PLKO.1-TRAF2-shR-NA(2),and PLKO.1-TRAF2-shRNA(3)lentivirus vector plasmids and control group lentivirus vector plasmids PLKO.1-puro were successfully constructed.The three TRAF2-shRNA lentivirus vector plasmids and control group lentivirus vector plasmids PLKO.1-puro were respectively introduced into the lentivirus packaging plasmid of HEK 293T to obtain virus solution.After infecting MH7A cells with the virus solution,they were treated with puromycin(2.00 μ G/mL)screening and obtaining MH7A stable transgenic plants after 2 days.Through qPCR and Western blot results,it was found that the expression of TRAF2 mRNA and protein in PLKO.1-TRAF2-shRNA(1)MH7A stably transfected cells was significantly reduced compared to the negative control group.The results of CCK-8 and Western blot showed that after knocking down TRAF2 in MH7A,the proliferation of MH7A cells with low TRAF2 expression induced by TNF-α and the phosphorylation level of P65 were significantly reduced.Conclusion A sta-ble transgenic strain of PLKO.1-TRAF2-shRNA(1)MH7A cells was successfully constructed to investigate the role of TNF-α-TRAF2 signal activation in mediating abnormal proliferation of RA synovial cells.

Objective:To analyze characteristics and trends of histopathological diagnosis of adult renal biopsy in Beijing from 2008 to 2020.Methods:A total of 4 652 cases of adult renal biopsy were collected from three hospitals in Beijing between 2008 and 2020. The patients were divided into three age groups: 18-40 years, 40-65 years and≥ 65 years; and also divided into three period: 2008-2011, 2012-2015, and 2016-2020. The pathological characteristics and changes of renal biopsy were analyzed in three age groups at different periods.Results:Among 4 652 cases primary glomerular disease accounted for 81.8%, the membranous nephropathy (MN, 32.4%, 1 509/4 652), IgA nephropathy (IgAN, 29.2%, 1 356/4 652) and minor glomerular abnormalities (MGA, 11.3%, 526/4 652) were the top three pathological types. The overall proportion of MN and diabetic nephropathy (DN) increased from 20.3% and 2.3% in 2008-2011 to 37.3% and 10.1% in 2016-2020) (χ2=99.9 and 96.1, both P<0.01), respectively. For age group 18-40 years, the MN and DN increased from 11.2% and 1.6% in 2008-2011 to 24.7% and 5.5% in 2016-2020 (χ2=32.7 and 20.7, both P<0.01), respectively. For age group 40-65 years the MN and DN increased from 26.6% and 3.2% in 2008-2011 to 41.5% and 13.1% in 2016-2020 (χ2=39.1 and 57.3, both P<0.01), respectively. For age group≥65 years the MN was the most common pathological type in the three periods, fluctuating between 41.3% and 55.0% (χ2=5.2, P=0.08); and DN increased from 0(0/63) in 2008-2011 to 7.5%(22/292) in 2016-2020 (χ2=8.1, P=0.02). Conclusion:The renal biopsy data show that membranous nephropathy and diabetic nephropathy are the most common primary and secondary adult glomerular diseases in Beijing recently.

Objective: To explore the relationship between dietary patterns and central precocious puberty in children, and to provide a scientific basis for dietary prevention of precocious puberty. Methods: A case-control study was conducted, among 35 newly diagnosed central precocious puberty girls from May to December 2019 as the case group, and 70 healthy girls with normal development as the control group. Physical development examination, parent questionnaire survey and child interview were carried out. Dietary information was assessed using a simplified food frequency questionnaire(FFQ). Principal component analysis was used to identify children s dietary patterns, and multiple Logistic regression was used to assess the association between dietary patterns and precocious puberty. Results: Three different dietary patterns have been established, namely "snack and processed food type", "animal protein type" and "nutritional tonic type" dietary patterns, respectively. After adjusting for covariates such as age and BMI, Logistic regression analysis showed that the "snack and processed food type" dietary pattern was positively correlated with precocious puberty(OR=10.81, 95%CI=2.59-45.15, P<0.01). There was a negative correlation between "animal protein type" and precocious puberty(OR=0.24, 95%CI=0.06-0.91, P=0.04), while the association between "nutritive tonic" and precocious puberty was not statistically significant(OR=0.28, 95%CI=0.07-1.05, P=0.06). Conclusion Children s dietary patterns were related to precocious puberty." Snack and processed food "dietary pattern with a high intake of fried foods, puffed foods, foods containing preservatives or pigments, western fast foods, chocolate and products, was closely related to central precocious puberty.

Objective: To explore the interaction effects and possible sex differences in childhood emotional overeating and polygenic influences on adolescent pubertal timing and tempo. Methods: In March 2016 (T0), all participants were recruited from grades 1 to 3 from two primary school of Bengbu, Anhui Province using cluster sampling, and follow up surveys were conducted once per year (T1, T2, T3). Emotional overeating was assessed at T1 and pubertal development was assessed annually (breast Tanner stage in girls and testicular volume in boys). The nonlinear growth model was used to estimate pubertal timing and tempo. Polygenic risk scores were calculated based on 17 SNPs for early pubertal timing. Hierarchical linear regression was performed to examine the interaction effects between childhood emotional overeating and polygenic risk scores on pubertal timing and tempo. Results: The complete data of 896 children were analyzed, including 373 boys (41.60%) and 523 girls (58.40%). A total of 203 (22.7%) children reported emotional overeating behavior at T1. After adjusting for several variables including early life adversity, delivery mode, and birthweight, only emotional overeating was associated with accelerated pubertal tempo among girls with a high genetic risk (B=0.19, 95%CI=0.07~0.32， P<0.01), although there was no association with pubertal timing (B=0.14, 95%CI=-0.12~0.41，P=0.28). In girls with a low genetic risk and boys, no evidence was found to support interaction effects between childhood emotional overeating and polygenic influences on pubertal timing and tempo (P＞0.05). Conclusion Emotional overeating was associated with a faster pubertal tempo in girls who had a high genetic risk of early pubertal development.

Objective: To evaluate the development trajectory of sugar-sweetened beverage (SSB) intake in childhood, and to explore the influence of different SSB intake patterns on childhood obesity. Methods: In 2016, a follow-up cohort study was carried out in two primary schools in Bengbu, Anhui Province. Three annual follow-ups were conducted in 1 263 children at baseline, and 997 children were included in the final analysis. Parental and student questionnaires were used to obtain basic information related to the children s consumption of SSBs. A group-based trajectory model (GBTM) was applied to classify the development trajectory of SSB intake patterns in childhood. Multiple linear regression analyses were performed to assess the correlation between different SSB intake patterns and childhood obesity. Results: GBTM identified four childhood SSB intake patterns, namely, the "persistently-low group (n=822), “decreasing-after-increasing” group (n=20), “gradually-decreasing” group (n=106), and “increasing” group (n=49). In the decreasing-after-increasing group and the gradually-decreasing group, baseline BMI levels and BMI levels obtained at the three follow-ups were significantly higher than those observed in the persistently-low group (F=6.26, 5.90, 5.99, 5.87, P<0.01). There were sex differences in the association between SSB intake patterns and the children s BMI levels. Among girls, after adjusting for confounding factors, the gradually decreasing group increased by 1.20 kg/m 2(B=1.20，95%CI=0.25-2.15, P=0.01) when compared with the persistently low group at the third follow-up. Among boys, no statistically significant association was found between SSB intake patterns and BMI levels (P>0.05). Conclusion Sex differences were observed with respect to the association between SSB intake patterns and obesity in children. Girls with a higher SSB intake had a significantly increased risk of obesity. Further studies are needed to explore the physiological mechanisms underlying sex differences, to provide the theoretical basis for developing intervention programs to prevent childhood obesity.

Objective:To investigate the role of neurite outgrowth inhibitor (Nogo)-oligodendrocyte myelin glycoprotein (Omgp)/Ras homologous (Rho)-Rho-associated coiled-coil forming protein kinase (Rock) signaling pathway in acute brain injury of carbon monoxide (CO) poisoning rats and treatment feasibility with Rho kinase inhibitor hydrochloride fasudil.Methods:According to random number table method, 135 healthy male SD rats were divided into three groups: a normal control group, a CO poisoning group and a fasudil treatment group ( n=45). Rat models of acute severe CO poisoning were established in the CO poisoning group and fasudil treatment group by inhalation method in a hyperbaric oxygen chamber. All rats received hyperbaric oxygen therapy for two weeks. Rats in the farsudil treatment group were intraperitoneally injected with hydrochloride farsudil for intervention (15 mg/[kg·d], once a d for 2 weeks), while those in the CO poisoning and normal control groups received the same volume of normal saline. The ultrastructures of rat brain tissues were observed by transmission electron microscopy one week after modeling. Staining intensities of Nogo- and OMgp-positive cells were detected by immunohistochemistry, and those of Rock-positive cells were analyzed by immunofluorescence one d, one week, one month and two months after modeling. The protein expressions of Nogo, OMgp and Rock in brain tissues were detected by Western blotting one d, one week, one month and two months after modeling. Results:In the CO poisoning group, the ultrastructures of brain tissues and blood-brain barrier were damaged obviously, and the changes in nucleus, mitochondria and synaptic structure were obvious; while fasudil treatment could effectively maintain the integrity of ultrastructures and functions of brain tissues, and reduce brain edema. One d, one week, one month and two months after modeling, the staining intensities of Nogo, OMgp and Rock positive cells and protein expression levels of Nogo, OMgp and Rock in the CO poisoning group were significantly higher than those in the normal control group at the same time point ( P<0.05); the staining intensities of Nogo, OMgp and Rock positive cells and protein expression levels of Nogo, OMgp and Rock in the fasudil treatment group were significantly lower than those in the CO poisoning group at the same time point ( P<0.05). Conclusion:The activation of Nogo-OMgp/Rho-Rock signaling pathway related molecules (Nogo, OMgp and Rock) is closely related to acute brain injury caused by CO poisoning; hydrochloride fasudil can effectively down-regulate the protein expressions of Nogo, OMgp and Rock, therefore obviously alleviate brain injury after CO poisoning.

BACKGROUND: Nano-hydroxyapatite/polyamide 66 (nHA/PA66) is a composite used widely in the repair of bone defects. However, this material is insufficient bioactivity. In contrast, D-RADA16-RGD self-assembling peptide (D-RADA16-RGD sequence containing all D-amino acids is Ac-RADARADARADARADARGDS-CONH2) shows admirable bioactivity for both cell culture and bone regeneration. Here, we describe the fabrication of a favorable biomaterial material (nHA/PA66/D-RADA16-RGD). METHODS: Proteinase K and circular dichroism spectroscopy were employed to test the stability and secondary structural properties of peptide D-RADA16-RGD respectively. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to characterize the surface of these materials. Confocal laser scanning (CLS), cell counting kit-8 tests (CCK-8), alizarin red S staining, cell immunofluorescence analysis and Western blotting were involved in vitro. Also biosafety and bioactivity of them have been evaluated in vivo. RESULTS: Proteinase K and circular dichroism spectroscopy demonstrated that D-RADA16-RGD in nHA/PA66 was able to form stable-sheet secondary structure. SEM and TEM showed that the D-RADA16-RGD material was 7–33 nm in width and 130–600 nm in length, and the interwoven pore size ranged from 40 to 200 nm. CLS suggests that cells in nHA/PA66/D-RADA16-RGD group were linked to adjacent cells with more actin filaments. CCK-8 analysis showed that nHA/PA66/D-RADA16-RGD revealed good biocompatibility. The results of Alizarin-red S staining and Western blotting as well as vivo osteogenesis suggest nHA/PA66/D-RADA16-RGD exhibits better bioactivity. CONCLUSION: This study demonstrates that our nHA/PA66/D-RADA16-RGD composite exhibits reasonable mechanical properties, biocompatibility and bioactivity with promotion of bone formation.
Actin Cytoskeleton ; Blotting, Western ; Bone Regeneration ; Cell Count ; Cell Culture Techniques ; Circular Dichroism ; Endopeptidase K ; Fluorescent Antibody Technique ; In Vitro Techniques ; Microscopy, Electron, Scanning ; Microscopy, Electron, Transmission ; Osteogenesis ; Sincalide ; Spectrum Analysis

The two-dimensional (2D) cell culture model is currently used to study cellular processes and drug screening for human diseases. However, the growth of cells is affected by many factors. For conventional 2D cell culture, many of the difficulties are encountered in accurately replicating the cell function in three-dimensional (3D) tissues. Compared with 2D cell culture, much attention is paid to the cell-to-cell and cell-matrix interactions for 3D cell culture systems, which can more closely mimic the growth environment for cultured cells. Therefore, the 3D cell culture system was more suitable for a variety of applications such as drug screening and cell proliferation. In this work, we prepared microarray-structured polymer films with different geometric structures by nanoimprint lithography and used the films as cell culture platforms for the culture of 293T cells. Through the adjustment of the surface morphology and water contact angle of the prepared films, the regulation of the morphological changes of cell growth was successfully realized. Experimental results demonstrated that the hydrophilic films with 10 μm-pillar microstructure are applicable to 3D cell culture, whereas the hydrophobic films with 3 μm-pillar microstructure are only suitable for 3D culture of cells with a smaller size and stiff cuticular layer. In addition, cells tended to the formation of spheroids on the hydrophobic films, while cells usually adhered to the surface and grew on the hydrophilic films. This work represents further technological progress in the development of 3D cell culture, thereby facilitating future studies of physiologically relevant processes.
Cell Culture Techniques ; Cell Proliferation ; HEK293 Cells ; Humans ; Polymers