Objective:To explore the regulation and mechanism of human placental extract(HPE)on lipopolysaccharide(LPS)-induced BV2 microglial cell inflammation.Methods:Microglia cell lines(BV2)were cultured in vitro and divided into PBS group,HPE group,LPS group and LPS+HPE group.BV2 cell viability was measured by CCK-8 analysis.A fluorescent probe targeting reactive oxygen species(ROS)was to detect the level of intracellular ROS.The mRNA levels of TNF-α,IL-1β and IL-6 were detected by RT-qPCR.The supernatants of different treatment groups were collected.The content of nitric oxide(NO)was detected by the Griess method,and the secretion levels of TNF-α,IL-1β and IL-6 were detected by the flow magnetic bead microarray(CBA)method.The indirect contact co-culture system between BV2 and mouse hippocampal neurons cell line HT22 cells was established to evaluate the neurotoxicity of HPE by the assessment of the cell viability and apoptosis of HT22 cells using CCK-8 or flow cytometry.The potential signaling molecules of NF-κB signaling pathway was detected by flow cytometry.Results:Compared with the PBS group,1.2 μg/ml LPS and 50 ng/ml HPE significantly inhibited the activity of BV2 cells.Compared with the PBS group,the mRNA levels of TNF-α,IL-1β and IL-6 in BV2 cells of the LPS group were significantly increased(P<0.05),and the secretion levels of NO,TNF-α,IL-1βand IL-6 in the supernatant were also significantly upregulated(P<0.05).The expressions of related signaling pathway molecules Toll-like receptor 4(TLR4),pIκBα and pNF-κB p65 were significantly increased(P<0.05).Compared with the LPS group,the mRNA levels of TNF-α,IL-1β and IL-6 in BV2 cells,the secretion levels of NO,TNF-α,IL-1β and IL-6,the neurotoxicity and neuronal apoptosis induced by microglial conditional medium,and the expressions of TLR4,pIκBα and pNF-κB p65 in the LPS+HPE group were significantly downregulate(P<0.05).Conclusion:HPE may alleviate the microglial inflammation,possibly through the TLR4/NF-κB signaling pathway.

Objective:To explore the regulation and mechanism of human placental extract(HPE)on lipopolysaccharide(LPS)-induced BV2 microglial cell inflammation.Methods:Microglia cell lines(BV2)were cultured in vitro and divided into PBS group,HPE group,LPS group and LPS+HPE group.BV2 cell viability was measured by CCK-8 analysis.A fluorescent probe targeting reactive oxygen species(ROS)was to detect the level of intracellular ROS.The mRNA levels of TNF-α,IL-1β and IL-6 were detected by RT-qPCR.The supernatants of different treatment groups were collected.The content of nitric oxide(NO)was detected by the Griess method,and the secretion levels of TNF-α,IL-1β and IL-6 were detected by the flow magnetic bead microarray(CBA)method.The indirect contact co-culture system between BV2 and mouse hippocampal neurons cell line HT22 cells was established to evaluate the neurotoxicity of HPE by the assessment of the cell viability and apoptosis of HT22 cells using CCK-8 or flow cytometry.The potential signaling molecules of NF-κB signaling pathway was detected by flow cytometry.Results:Compared with the PBS group,1.2 μg/ml LPS and 50 ng/ml HPE significantly inhibited the activity of BV2 cells.Compared with the PBS group,the mRNA levels of TNF-α,IL-1β and IL-6 in BV2 cells of the LPS group were significantly increased(P<0.05),and the secretion levels of NO,TNF-α,IL-1βand IL-6 in the supernatant were also significantly upregulated(P<0.05).The expressions of related signaling pathway molecules Toll-like receptor 4(TLR4),pIκBα and pNF-κB p65 were significantly increased(P<0.05).Compared with the LPS group,the mRNA levels of TNF-α,IL-1β and IL-6 in BV2 cells,the secretion levels of NO,TNF-α,IL-1β and IL-6,the neurotoxicity and neuronal apoptosis induced by microglial conditional medium,and the expressions of TLR4,pIκBα and pNF-κB p65 in the LPS+HPE group were significantly downregulate(P<0.05).Conclusion:HPE may alleviate the microglial inflammation,possibly through the TLR4/NF-κB signaling pathway.

Objective To investigate the potential of a two-component model combining T2 and diffusion weighted imaging(T2-DWI)in the diagnosis of prostate cancer.Methods Thirty-two prostate cancer patients and twenty prostatic hyperplasia patients underwent combined T2-DWI,with TE values set at 50 ms and 70 ms and b values at 50 s/mm2 and 800 s/mm2,respectively.This provided four measurements for each voxel.The signal fractions were calculated from the slow component(SFslow)extracted from the two-component model of T2-DWI,which was fitted with two free parameters.The results were compared with the traditional single exponential apparent diffusion coefficient(ADC)model.Results A significant difference was observed between SFslow(0.77±0.14 vs 0.35±0.10)and ADC[(0.91±0.24)μm2/ms vs(1.98±0.32)μm2/ms]in region of interest(ROI)between peripheral zone tumors and normal prostate tissue(P＜0.001).No significant difference was found between SFslow(0.75±0.13 vs 0.68±0.11)and ADC[(0.88±0.18)μm2/ms vs(0.97±0.13)μm2/ms]in ROI between non-peripheral zone tumors and prostatic hyperplasia.The area under the curve(AUC)of the receiver operating characteristic(ROC)for distinguishing tumors from prostate voxels was 0.962 for two-component SFslow and 0.940 for single exponential ADC models.The Spearman correlation coefficients between tumor SFslow and ADC with Gleason scores were 0.631 and-0.558,respectively.Conclusion SF estimation based on T2-DWI two-component model can effectively differentiate tumors from normal prostate tissue,showing potential in diagnosing prostate cancer.

Objective:To investigate the effect of combination of infrared thermography (IRT) and high frequency colour Doppler ultrasound (HFCDU) in location of superficial fascia perforating vessels and to guide the design and harvest of super-thin anterolateral thigh flap (ALTF) .Methods:A total of 15 patients who received medical treatment in the Department of Orthopaedics, Zhoushan Hospital and Department of Foot and Ankle Surgery, Wuxi No.9 People’s Hospital from January 2022 to July 2023 were selected to participate the study. The patients were 11 males and 4 females, aged 26 to 64 years with an average age of 48.3 years. A total of 14 wounds of foot, 1 of hand, 1 of forearm and 1 of ankle were reconstructed with 17 free super-thin ALTFs. The sizes of soft tissue defect were 5 cm×3 cm-23 cm×7 cm. The flaps were 6 cm×4 cm to 25 cm×8 cm in size. The donor sites were directly pulled together and sutured. Before surgery, HFCDU was applied to locate the perforator vessels across deep fascia into superficial fascia of the ALTF. IRT was further employed to locate the superficial perforator vessels on the superficial fascia in operating room. The running course of a perforating branch in the superficial fascia was determined by the running courses of the perforating branch located by the two location methods. Super-thin ALTFs were harvested and the precise locations of the perforating vessels were verified in surgery. SPSS 23.0 software was used for data analysis. Rank sum test was performed on the location data of superficial fascial perforating branches found by IRT and the data in surgery. P<0.05 was considered a statistically significant difference. All patients were included in the postoperative follow-up by means of visit of outpatient clinic and via WeChat reviews, where the survival and functional recovery of the flaps were observed. Results:A total of 30 perforating vessels had been located by HFCDU and 30 by IRT, and a total of 31 perforating vessels were found in superficial fascia in surgery. The true positive rate was 93.3%, with a false positive rate at 6.7% and a false negative rate at 9.7%. The rank sum test calculated P=0.853 for the number of perforating vessels located by the IRT and those found from surgery. There was no significant difference between the 2 detecting methods. No postoperative complication occurred in all 14 flaps. Partial necrosis occurred in 1 flap but healed after dressing changes. Venous occlusion had occurred in 1 flap, it was rectified after surgical exploration. Superficial infection happened in 1 flap and it was improved after anti-infection treatment. Postoperative follow-up was conducted for 3-12 months. The flaps were in good texture with satisfactory appearance and function of limbs. All donor sites healed well without scar hyperplasia. Conclusion:IRT combined with HFCDU is a reliable method in location of perforator vessels of ALTF, and it is an ideal technique in the exploration of perforator vessels and in the harvest of a super-thin flap.

Objective:To investigate the effects and mechanisms of exogenous calcium-binding protein S100A4 on prolifera-tion,survival and migration functions of glioma cells.Methods:Pan-cancer dataset was downloaded from UCSC database for the analysis of S100A4 expression and prognosis in pan-cancer,and transcriptome and clinical data of glioma patients were downloaded from CGGA database.Prognosis and progression of S100A4 expression in glioma patients were analyzed by R software.S100A4 protein network interaction of glioma patients were addressed by online analytical tools GEPIA and STRING.Human glioma cell lines(U87 and U251)were cultured in vitro,and the experiment was divided into 3 groups:PBS group,S100A4 group and S100A4+TAK242 group[Resa-torvid(TAK242)was a specific inhibitor of Toll-like receptors 4(TLR4)].Flow cytometry was used to detect proliferation and apopto-sis of glioma cells.Wound healing assay,Transwell assay and clone formation assay were used to detect migration and proliferation ability of U87 and U251 cells,and Western blot was used to detect levels of TLR4 protein and NF-κB related signaling proteins.Results:Bioinformatics results showed that S100A4 was significantly upregulated in multiple tumours(P<0.05),which included glio-blastoma(GBM),lower grade glioma(LGG),stomach adenocarcinoma(STAD),liver hepatocellular carcinoma(LIHC),etc,with poorer prognosis in GBM and LGG.Compared with glioma patients with low expression of S1004,high expression S100A4 in glioma patients had shorter survival and higher degree of WHO classification.In addition,S100A4 protein in glioma patients may interact with Annexin A2(ANXA2),TLR4 and advanced glycosylation end product-specific receptor(AGER)proteins.In cellular experiments,U87 and U251 cells showed enhanced proliferation and migration,and significantly up-regulated levels of TLR4,p-ERK1/2,p-p38 and p-p65 proteins after exogenous S100A4 treatment compared to PBS group(P<0.05),while glioma cells in S100A4+TAK242 group showed weaker proliferation and migration,and lower TLR4,p-p38 and p-p65 protein levels than those in S100A4 group,TLR4,p-ERK1/2,p-p38,p-p65 protein levels were significantly down-regulated(P<0.05).Conclusion:S100A4 may regulate func-tion of glioma proliferation,migration and survival through TLR4/NF-κB signaling pathway.

Objective:To investigate the effect of combination of infrared thermography (IRT) and high frequency colour Doppler ultrasound (HFCDU) in location of superficial fascia perforating vessels and to guide the design and harvest of super-thin anterolateral thigh flap (ALTF) .Methods:A total of 15 patients who received medical treatment in the Department of Orthopaedics, Zhoushan Hospital and Department of Foot and Ankle Surgery, Wuxi No.9 People’s Hospital from January 2022 to July 2023 were selected to participate the study. The patients were 11 males and 4 females, aged 26 to 64 years with an average age of 48.3 years. A total of 14 wounds of foot, 1 of hand, 1 of forearm and 1 of ankle were reconstructed with 17 free super-thin ALTFs. The sizes of soft tissue defect were 5 cm×3 cm-23 cm×7 cm. The flaps were 6 cm×4 cm to 25 cm×8 cm in size. The donor sites were directly pulled together and sutured. Before surgery, HFCDU was applied to locate the perforator vessels across deep fascia into superficial fascia of the ALTF. IRT was further employed to locate the superficial perforator vessels on the superficial fascia in operating room. The running course of a perforating branch in the superficial fascia was determined by the running courses of the perforating branch located by the two location methods. Super-thin ALTFs were harvested and the precise locations of the perforating vessels were verified in surgery. SPSS 23.0 software was used for data analysis. Rank sum test was performed on the location data of superficial fascial perforating branches found by IRT and the data in surgery. P<0.05 was considered a statistically significant difference. All patients were included in the postoperative follow-up by means of visit of outpatient clinic and via WeChat reviews, where the survival and functional recovery of the flaps were observed. Results:A total of 30 perforating vessels had been located by HFCDU and 30 by IRT, and a total of 31 perforating vessels were found in superficial fascia in surgery. The true positive rate was 93.3%, with a false positive rate at 6.7% and a false negative rate at 9.7%. The rank sum test calculated P=0.853 for the number of perforating vessels located by the IRT and those found from surgery. There was no significant difference between the 2 detecting methods. No postoperative complication occurred in all 14 flaps. Partial necrosis occurred in 1 flap but healed after dressing changes. Venous occlusion had occurred in 1 flap, it was rectified after surgical exploration. Superficial infection happened in 1 flap and it was improved after anti-infection treatment. Postoperative follow-up was conducted for 3-12 months. The flaps were in good texture with satisfactory appearance and function of limbs. All donor sites healed well without scar hyperplasia. Conclusion:IRT combined with HFCDU is a reliable method in location of perforator vessels of ALTF, and it is an ideal technique in the exploration of perforator vessels and in the harvest of a super-thin flap.

Objective:To explore the association between hair trace element and all-cause death in the elderly in Hainan Province.Methods:The subjects of the study were elderly people from China Hainan Centenarian Cohort Study, a total of 163 elderly were included. The association between hair trace element level and all-cause death was analyzed by using Cox proportional risk regression model.Results:After fully adjusting the covariates, the multiple Cox proportional hazards regression analyses showed that selenium (Se), manganese (Mn), strontium (Sr) concentrations in hair were significantly associated with all-cause mortality, the hazard ratio ( HR) were 0.72 (95% CI: 0.54-0.98, P=0.035), 1.50 (95% CI: 1.07-2.11, P=0.020) and 0.54 (95% CI: 0.37-0.79, P=0.001), respectively. Subgroup and cross analysis showed that hair copper (Cu) were significant association with death in the people with anemia, the HR were 1.81 (95% CI: 1.13-2.88, P=0.013). And, hair Mn interacted with anemia, the HR was 0.46 (95% CI: 0.22-0.94, P=0.033). Conclusions:Se, Mn and Sr concentrations in hair were associated with the elevated risk for all-cause death in the elderly in Hainan. Se, Mn and Sr concentrations in hair can be used as a reference index for the prediction of the death risk of long-lived elderly in community, suggesting that the daily diet of elderly people are rich and diverse, in order to maintain normal and balanced trace element content in the body.

OBJECTIVE: To investigate the effect of Zuogui Jiangtang Jieyu Decoction (ZJJ) on Shh signaling and self-renewal of neural stem cells in the hippocampal dentate gyrus of diabetic rats with depression. METHODS: Diabetic rat models with depression were randomly divided into model group, positive drug (metformin + fluoxetine) group, and low-, medium-, and high-dose ZJJ groups (n=16), with normal SD rats as the control group. The positive drugs and ZJJ were administered by gavage, and the rats in the control and model groups were given distilled water. After the treatment, blood glucose level was detected using test strips, and behavioral changes of the rats were assessed by forced swimming test and water maze test. ELISA was used to examine the serum level of leptin; The expressions of nestin and Brdu proteins in the dentate gyrus of the rats were detected using immunofluorescence assay, and the expressions of self-renewal marker proteins and Shh signaling proteins were detected using Western blotting. RESULTS: The diabetic rats with depression showed significantly increased levels of blood glucose and leptin (P < 0.01) and prolonged immobility time in forced swimming test (P < 0.01) and increased stage climbing time with reduced stage seeking time and stage crossings in water maze test (P < 0.01). The expressions of nestin and Brdu in the dentate gyrus, the expressions of cyclin D1, SOX2, Shh, Ptch1, Smo in the hippocampus and the nuclear expression of Gli-1 were decreased (P < 0.01) while hippocampal Gli-3 expression was increased significantly (P < 0.01) in the rat models. Treatment of rat models with high-dose ZJJ significantly reduced the blood glucose (P < 0.01) and leptin level (P < 0.05) and improved their performance in behavioral tests (P < 0.01). The treatment also obviously increased the expressions of nestin, Brdu, cyclin D1, SOX2, Shh, Ptch1, and Smo and the nuclear expression of Gli-1 in the dentate gyrus (P < 0.01) and reduced hippocampal expression of Gli-3 (P < 0.05) in the rat models. CONCLUSION ZJJ can significantly improve the self-renewal ability of neural stem cells and activate Shh signaling in dentate gyrus of diabetic rats with depression.
Animals ; Rats ; Blood Glucose ; Bromodeoxyuridine ; Cell Self Renewal ; Cyclin D1 ; Dentate Gyrus ; Depression ; Diabetes Mellitus, Experimental ; Hippocampus ; Leptin ; Nestin ; Rats, Sprague-Dawley

Cadmium (Cd) is a common heavy metal in the environment. Cd²⁺ may penetrate the blood-brain barrier and produce neurotoxicity, thus inducing various neurodegenerative diseases. Celastrol is an effective component of Tripterygium wilfordii Hook. F., which has many pharmacological effects such as anti-cancer and anti-inflammatory. Here we explored the effect of celastrol on the corresponding neurotoxicity induced by Cd²⁺. Cell proliferation test, cell membrane integrity test, and cell morphology were observed to analyze the effect of Cd²⁺ on the viability of HMC3. The neurotoxicity of Cd²⁺ and the effect of celastrol on the corresponding neurotoxicity induced by Cd²⁺ were analyzed by nitric oxide (NO) test, lipid peroxidation (MDA) test, and Western blotting. When the concentration of Cd²⁺ reached 40 μmol/L, the inhibition rate of HMC3 cell proliferation was (57.17±8.23)% (P < 0.01, n=5), compared with the control group. The cell activity continued to reduce when the Cd²⁺ concentration further increased. When the concentration of Cd²⁺ was higher than 40 μmol/L, the cell membrane of HMC3 was significantly damaged, and the damage was dose-dependent. Upon increasing the Cd²⁺ concentration, the cell morphology began to change and the adhesion also became worse. Cd²⁺ significantly increased the amount of NO released by HMC3 cells, while celastrol effectively inhibited the NO release of HMC3 cells induced by Cd²⁺. Cd²⁺ greatly increased the release of MDA in HMC3 cells, and the level of MDA decreased rapidly upon the addition of 10^-7 mol/L celastrol. Cd²⁺ increased the expression of p-PI3K protein, and the levels of p-PI3K protein and p-AKT protein were inhibited by the addition of celastrol (10^‒7 mol/L, 10^‒6 mol/L), thus preventing cell apoptosis. In conclusion, celastrol inhibits Cd²⁺ induced microglial cytotoxicity and plays a neuroprotective role.
Anti-Inflammatory Agents/pharmacology* ; Apoptosis ; Cadmium/toxicity* ; Nitric Oxide/pharmacology* ; Pentacyclic Triterpenes ; Phosphatidylinositol 3-Kinases ; Proto-Oncogene Proteins c-akt/metabolism* ; Triterpenes/pharmacology*

The development and implement of microbial chassis cells can provide excellent cell factories for diverse industrial applications, which help achieve the goal of environmental protection and sustainable bioeconomy. The synthetic biology strategy of Design-Build-Test-Learn (DBTL) plays a crucial role on rational and/or semi-rational construction or modification of chassis cells to achieve the goals of "Building to Understand" and "Building for Applications". In this review, we briefly comment on the technical development of the DBTL cycle and the research progress of a few model microorganisms. We mainly focuse on non-model bacterial cell factories with potential industrial applications, which possess unique physiological and biochemical characteristics, capabilities of utilizing one-carbon compounds or of producing platform compounds efficiently. We also propose strategies for the efficient and effective construction and application of synthetic microbial cell factories securely in the synthetic biology era, which are to discover and integrate the advantages of model and non-model industrial microorganisms, to develop and deploy intelligent automated equipment for cost-effective high-throughput screening and characterization of chassis cells as well as big-data platforms for storing, retrieving, analyzing, simulating, integrating, and visualizing omics datasets at both molecular and phenotypic levels, so that we can build both high-quality digital cell models and optimized chassis cells to guide the rational design and construction of microbial cell factories for diverse industrial applications.
Bacteria/genetics* ; Metabolic Engineering ; Synthetic Biology