AIM: To investigate the mechanism by which Yijing Decoction antagonist high glucose-induced damage to the basement membrane(BM)in an in vitro inner blood-retinal barrier(iBRB)model.METHODS:Rat retinal microvascular pericytes(RMPs)and endothelial cells(ECs)were isolated and cultured to establish an in vitro iBRB model. The cells were randomly divided into four groups: low glucose group(LG), high glucose group(HG), minocycline group(MG)and Yijing Decoction group(YG). The LG group received 25 mmol/L glucose, the HG group received 60 mmol/L glucose, the MG group received 60 mmol/L glucose + 10 μg/mL minocycline, and the YG group received 60 mmol/L glucose + 10% Yijing Decoction-containing serum. Incubation for each group were terminated after intervention for 12 h. Next, the Western blot analysis was performed to assess the protein expression of BM-related proteins, including collagen Ⅳ(CⅣ)and laminin(LN), as well as matrix metalloproteinase(MMPs)/tissue inhibitor of matrix metalloproteinases(TIMPs)such as MMP-2, MMP-3, MMP-9, TIMP-1, TIMP-2.RESULTS:Compared to the LG group, the protein expressions of CⅣ increased in the HG, MG, and YG groups, as did LN in the HG and MG groups(all P<0.05). Both Yijing Decoction and minocycline effectively inhibited the elevated expression of CⅣ and LN induced by high glucose, and the difference between the YG, MG, and HG groups was statistically significant(all P<0.05). Futhermore, compared to the LG group, the protein expressions of MMP-2, MMP-3, and MMP-9 increased in the HG, MG, and YG groups(all P<0.05). Yijing Decoction specifically attenuated the high glucose-induced increase in MMP-2, MMP-3 and MMP-9 protein expression, and there were statistically significant differences between the YG and HG group(all P<0.05). No significant difference were observed in the expressions of TIMP-1 and TIMP-2 among the LG, HG, MG, and YG groups(all P>0.05).CONCLUSION:Yijing Decoction can potentially intervene in DR by modulating the protein expression of MMP-2, MMP-3, MMP-9, CⅣ, and LN, suppressing high glucose-induced BM remodeling, and mitigating damage to iBRB.

Background::Bladder cancer, characterized by a high potential of tumor recurrence, has high lifelong monitoring and treatment costs. To date, tumor cells with intrinsic softness have been identified to function as cancer stem cells in several cancer types. Nonetheless, the existence of soft tumor cells in bladder tumors remains elusive. Thus, our study aimed to develop a microbarrier microfluidic chip to efficiently isolate deformable tumor cells from distinct types of bladder cancer cells.Methods::The stiffness of bladder cancer cells was determined by atomic force microscopy (AFM). The modified microfluidic chip was utilized to separate soft cells, and the 3D Matrigel culture system was to maintain the softness of tumor cells. Expression patterns of integrin β8 (ITGB8), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) were determined by Western blotting. Double immunostaining was conducted to examine the interaction between F-actin and tripartite motif containing 59 (TRIM59). The stem-cell-like characteristics of soft cells were explored by colony formation assay and in vivo studies upon xenografted tumor models. Results::Using our newly designed microfluidic approach, we identified a small fraction of soft tumor cells in bladder cancer cells. More importantly, the existence of soft tumor cells was confirmed in clinical human bladder cancer specimens, in which the number of soft tumor cells was associated with tumor relapse. Furthermore, we demonstrated that the biomechanical stimuli arising from 3D Matrigel activated the F-actin/ITGB8/TRIM59/AKT/mTOR/glycolysis pathways to enhance the softness and tumorigenic capacity of tumor cells. Simultaneously, we detected a remarkable up-regulation in ITGB8, TRIM59, and phospho-AKT in clinical bladder recurrent tumors compared with their non-recurrent counterparts.Conclusions::The ITGB8/TRIM59/AKT/mTOR/glycolysis axis plays a crucial role in modulating tumor softness and stemness. Meanwhile, the soft tumor cells become more sensitive to chemotherapy after stiffening, that offers new insights for hampering tumor progression and recurrence.

Arrhythmogenic cardiomyopathy (ACM), a fatal heart disease characterized by fibroadipocytic replacement of cardiac myocytes, accounts for 20% of sudden cardiac death and lacks effective treatment. It is often caused by mutations in desmosome proteins, with Desmoglein-2 (DSG2) mutations as a common etiology. However, the mechanism underlying the accumulation of fibrofatty in ACM remains unknown, which impedes the development of curative treatment. Here we investigated the fat accumulation and the underlying mechanism in a mouse model of ACM induced by cardiac-specific knockout of Dsg2 (CS-Dsg2 ^-/-). Heart failure and cardiac lipid accumulation were observed in CS-Dsg2 ^-/- mice. We demonstrated that these phenotypes were caused by decline of fatty acid (FA) β-oxidation resulted from impaired mammalian target of rapamycin (mTOR) signaling. Rapamycin worsened while overexpression of mTOR and 4EBP1 rescued the FA β-oxidation pathway in CS-Dsg2 ^-/- mice. Reactivation of PPARα by fenofibrate or AAV9-Pparα significantly alleviated the lipid accumulation and restored cardiac function. Our results suggest that impaired mTOR-4EBP1-PPARα-dependent FA β-oxidation contributes to myocardial lipid accumulation in ACM and PPARα may be a potential target for curative treatment of ACM.

Background::DOK3 (Downstream of kinase 3) is involved primarily with immune cell infiltration. Recent research reported the role of DOK3 in tumor progression, with opposite effects in lung cancer and gliomas; however, its role in prostate cancer (PCa) remains elusive. This study aimed to explore the role of DOK3 in PCa and to determine the mechanisms involved.Methods::To investigate the functions and mechanisms of DOK3 in PCa, we performed bioinformatic and biofunctional analyses. Samples from patients with PCa were collected from West China Hospital, and 46 were selected for the final correlation analysis. A lentivirus-based short hairpin ribonucleic acid (shRNA) carrier was established for silencing DOK3. A series of experiments involving the cell counting kit-8, bromodeoxyuridine, and flow cytometry assays were performed to identify cell proliferation and apoptosis. Changes in biomarkers from the nuclear factor kappa B (NF-κB) signaling pathway were detected to verify the relationship between DOK3 and the NF-κB pathway. A subcutaneous xenograft mouse model was performed to examine phenotypes after knocking down DOK3 in vivo. Rescue experiments with DOK3 knockdown and NF-κB pathway activation were designed to verify regulating effects. Results::DOK3 was up-regulated in PCa cell lines and tissues. In addition, a high level of DOK3 was predictive of higher pathological stages and worse prognoses. Similar results were observed with PCa patient samples. After silencing DOK3 in PCa cell lines 22RV1 and PC3, cell proliferation was significantly inhibited while apoptosis was promoted. Gene set enrichment analysis revealed that DOK3 function was enriched in the NF-κB pathway. Mechanism experiments determined that knockdown of DOK3 suppressed activation of the NF-κB pathway, increased the expressions of B-cell lymphoma-2 like 11 (BIM) and B-cell lymphoma-2 associated X (BAX), and decreased the expression of phosphorylated-P65 and X-linked inhibitor of apoptosis (XIAP). In the rescue experiments, pharmacological activation of NF-κB by tumor necrosis factor-α (TNF-α) partially recovered cell proliferation after the knockdown of DOK3.Conclusion::Our findings suggest that overexpression of DOK3 promotes PCa progression by activating the NF-κB signaling pathway.

Objective:To explore the difference in neurotransmitter levels between persons who had been comatose after a brain injury and healthy counterparts, and to explore the relationship between neurotransmitters and different states of consciousness.Methods:A total of 52 persons with brain injury were analyzed retrospectively and divided into a coma group of 30 with Glasgow coma scale scores ≤9 and a sober group of 22 with scores >9. γ-GABA, 5-hydroxytryptamine, noradrenaline and dopamine levels were measured using encephalofluctuography and compared between the two groups.Results:The levels of all of the neurotransmitters tested were significantly lower in the coma group than in the sober group. There were, however, no significant differences between the two groups in terms of relative levels of the neurorsmters. Logistic regression showed that dopamine deficiency is an important risk factor for coma after a brain injury.Conclusions:Most neurotransmitters decrease significantly among those comatose after a brain injury. Dopamine deficiency seems to be a risk factor for the incidence of coma. Therefore, drugs or rehabilitation therapies which can improve dopamine levels can be considered to help coma patients regain consciousness.

Objective To investigate the role of microRNA-1 (miR-1) in cardiac fibroblasts induced by high glucose in rats. Methods The primary fibroblasts were cultured from the apical tissue of 1-3 day-old Sprague-Dawley (SD) rats. The cells which were passaged to generation 3 or 4, were randomly divided into normal glucose+lentivector-vehicle group (CON+Lv-Vehicle group), normal glucose+lentivector-miR-1 group (CON+Lv-miR1 group), high glucose+lentivector-vehicle group (HG+Lv-Vehicle group), high glucose+lentivector-miR-1 group (HG+Lv-miR1 group), high glucose+Lv-Vehicle+inhibitor group (HG+Lv-Vehicle+CC group), and high glucose+lentivector-miR-1+inhibitor group (HG+Lv-miR1+CC group). The myocardial fibroblasts were cultured in the concentration of 5.5 mmol/L glucose (normal glucose) or 25.0 mmol/L (high glucose) DMEM medium. Then lentiviral vector containing miR-1 silent sequence or the same volume of lentiviral vector was inoculated into the cells. The AMP activated protein kinase (AMPK) inhibitor Compound C (20 μmol/L) was added to the medium at 12 hours before sampling in inhibitor groups. The expression of phosphorylation of AMPK (p-AMPK), collagenⅠandⅢ, matrix metalloproteinase (MMP-2, MMP-9), and autophagy flux related protein LC3B-Ⅱ and p62/SQSTM1 were measured by Western Blot. Results The purity of rat myocardial fibroblasts in vitro was 97%. Compared with CON+Lv-Vehicle group, there was no significant difference in the expression of p-AMPK in CON+Lv-miR1 group, the expression of p-AMPK in HG+Lv-Vehicle group was significantly decreased (p-AMPK/t-AMPK: 44.72±3.29 vs. 100.00±7.77, 1 < 0.01). The expression of p-AMPK in HG+Lv-miR1 group was higher than that in HG+Lv-Vehicle group (p-AMPK/t-AMPK:60.52±5.16 vs. 44.72±3.29, 1 < 0.05). Compared with HG+Lv-Vehicle group, the expressions of collagen, MMP, LC3B-Ⅱand p62/SQSTM1 in HG+Lv-miR1 group were significantly decreased; after the treatment with AMPK inhibitor, the expressions of collagen, MMP, LC3B-Ⅱ, p62/SQSTM1 were significantly increased (HG+Lv-Vehicle+CC group vs. HG+Lv-Vehicle group: collagen Ⅰ/β-actin: 158.74±13.21 vs. 100.00±7.64, collagenⅢ/β-actin: 177.38± 17.31 vs. 100.00±5.18, MMP-2/β-actin: 130.09±14.31 vs. 100.00±10.47, MMP-9/β-actin: 215.54±20.92 vs. 100.00±11.28, LC3B-Ⅱ/β-actin: 159.34±13.83 vs. 100.00±6.44, p62/SQSTM1/β-actin: 201.01±24.02 vs. 100.00±8.62; HG+Lv-miR1+CC group vs. HG+Lv-miR1 group: collagenⅠ/β-actin: 108.69±9.93 vs. 80.83±7.24, collagenⅢ/β-actin: 127.68±10.46 vs. 81.56±9.97, MMP-2/β-actin: 106.66±10.21 vs. 74.80±7.43, MMP-9/β-actin: 145.65±11.56 vs. 74.63±10.55, LC3B-Ⅱ/β-actin: 150.15±13.28 vs. 22.98±2.87, p62/SQSTM1/β-actin: 130.48±10.74 vs. 49.90±2.27, all 1 < 0.05). Conclusion miR-1 gene silencing inhibits myocardial fibrosis induced by high glucose, its mechanism may be related to the up-regulation of p-AMPK, which can recover autophagy flux.

Objective:to apply Fuzhengkangai AIDS particles combined therapy with effective antiretroviral therapy (HAART) at the beginning of the HIV/AIDS (human immunodeficiency virus/acquired immunodeficiency syndrome) patients,observe its clinical curative effect and improve the related adverse reactions.Methods:a multicenter,randomized,double-blind,placebo-controlled clinical research method,divided into experimental group and the control group (n =57),experimental group to "Fuzhengkangai particles" joint efficient antiretroviral therapy (HAART) antiviral treatment,control group to "Fuzhengkangai particles" placebo (5％ low dose) joint efficient antiretroviral therapy (HAART) antiviral treatment,treatment for 3 months.Two groups of patients with simultaneous liver and kidney function,blood routine,CD4 + T lymphocyte count,the observation of TCM symptoms and signs,survival quality of integral.Results:after 12 weeks of treatment,the control group and experimental group,the experimental group significantly elevated levels of peripheral blood RBC,lower levels of CR,reduce the signs and symptoms (total score,fatigue,stay,and spontaneous perspiration)integral,higher levels of CD4 + T lymphocyte count and survival quality score,the difference had statistical significance (p ＜ 0.05).Conclusion:Fuzhengkangai particles joint efficient antiretroviral therapy (HAART) antiviral treatment,can improve HAART after initial treatment of the patient's clinical symptoms,blood toxicity,the quality of survival,reduce the antiviral treatment of adverse reactions,combination drug alone can significantly increase the CD4 + T lymphocytes,so as to improve clinical curative effect.

OBJECTIVETo analyze the clinical feature and constituent ratio of adult hip fractures in Southwest China.

METHODSThe data of adult inpatients and outpatients with hip fractures treated between January 2010 and December 2011 in 11 hospitals of the Southwest China were collected and analyzed. The data includes gender, age, age distribution and fracture pattern according to AO classification.

RESULTSThere were a total of 2,833 adult hip fractures, including 1,340 (47.30%) males and 1,493 (52.70%) females, with a male-to-female incidence ratio of 1: 1.11 and a mean age of (66±18) years. The highest frequency of hip fractures was seen in the 71 to 85 years age group (42.18%, 1,195/2,833). There were 844 fractures (29.79%) in the young and middle-aged group (16-<60 years) and 1 898 fractures (70.21%) in the geriatric group (≥60 years). Men had a higher rate than women (men: 577 fractures, 68.4%) in the young and middle-aged group, while women had a higher rate than men (women: 1,226 fractures, 61.64%) in the geriatric group, with a significant difference in the sex distribution between the two groups (χ2=214.001, P<0.01). The proportion of intertrochanteric fracture (type 31-A), femoral neck (type 31-B) and femoral head fracture (type 31-C) was 46.59%, 49.74% and 3.67% respectively. The highest frequency of the sub-type in each fracture type was type 31-A2, type 31-B2 and type 31-C2.

CONCLUSIONSWomen have a higher rate than men in Southwest China. Geriatric patients are more than the young and middle-aged patients. The femoral neck fractures, intertrochanteric fractures and femoral head fractures are in descending orders according to the proportion of the three different hip fractures.

Aged ; Aged, 80 and over ; China ; epidemiology ; Female ; Femoral Fractures ; Femoral Neck Fractures ; Femur ; Femur Head ; Femur Neck ; Hip Fractures ; epidemiology ; Humans ; Incidence ; Male ; Middle Aged

BACKGROUND:Unlike the ilium derived from the paraxial mesoderm, the mandible from cranial neural crest has a unique mechanism. Core binding factorα1 (Cbfα1) is a key transcription factor for skeletogenic process. However, the role of Cbfα1/p56 subtype in mandible tissue is yet not clear. OBJECTIVE:To research the expression of Cbfα1/p56 subtype in bone marrow mesenchymal cells from rat mandible in vitro. METHODS:Bone marrow mesenchymal stem cells from rat mandible and ilium were in vitro isolated and purified by primary culture. The characteristics of bone marrow mesenchymal cells were compared through the methods of enzyme linked immunosorbent assay and real-time PCR, including growth curve, alkaline phosphatase activity and relative mRNA expression of Cbfα1 subtypes. RESULTS AND CONCLUSION:Bone marrow mesenchymal cells from rat mandible and ilium were successful y obtained. Bone marrow mesenchymal cells from the mandible proliferated more rapidly, alkaline phosphatase activity of which was higher than iliac cells. The relative mRNA expression of Cbfα1/p56 subtype in bone marrow mesenchymal cells from the mandible was more than that in iliac cells at 6 days of culture (P<0.05), while the expression of Cbfα1/p57 in each time showed no statistical significance (P>0.05). The results showed that Cbfα1/p56 is very significant in the early osteogenic differentiation of bone marrow mesenchymal cells from the mandible.

A robust,selective and highly sensitive chemiluminescent (CL) platform for protein assay was presented in this paper.This novel CL approach utilized rolling circle amplification (RCA) as a signal enhancement technique and the 96-well plate as the immobilization and separation carrier.Typically,the antibody immobilized on the surface of 96-well plate was sandwiched with the protein target and the aptamer-primer sequence.This aptamer-primer sequence was then employed as the primer of RCA.Based on this design,a number of the biotinylated probes and streptavidin-horseradish peroxidase (SA-HRP) were captured on the plate,and the CL signal was amplified.In summary,our results demonstrated a robust biosensor with a detection limit of 10 fM that is easy to be established and utilized,and devoid of light source.Therefore,this new technique will broaden the perspective for future development of DNA-based biosensors for the detection of other protein biomarkers related to clinical diseases,by taking advantages of high sensitivity and selectivity.