Objective To construct HEK 293T cells that express tardigrade Dsup protein fused with green fluorescent protein copGFP in order to study the effect of Dsup protein on proliferation of HEK 293T cells.Methods The CRISPR/Cas9 gene knock-in system was constructed.The target gene fragments of Dsup,copGFP,EF1α and puromycin were amplified by PCR and inserted into pAAVS1-SFFV to construct the fusion vector of Dsup and copGFP,which was known as pAAVS1-SFFV-Dsup-copGFP-EF1α-Puro.pAAVS1-SFFV-Dsup-copGFP-EF1 α-Puro and pAAVS1-CRISPR-Cas9 vector were co-transfected into HEK 293T cells before Dsup gene was inserted into the AAVS1 region of HEK 293T cells via homologous recombination.The HEK 293T cells expressing Dsup gene were obtained following puromycin selection,flow cytometry sorting and genome identification.The expression of Dsup at mRNA and protein levels and proliferation-related genes(MCM2,MCM4,PCNA,Ki-67)were examined to investigate the effects of Dsup gene on the proliferation of HEK 293T-Dsup-copGFP cells.Results The pAAVS1-SFFV-Dsup-copGFP-EF1α-Puro recombinant vector was constructed,and the HEK 293T-Dsup-copGFP cells with Dsup gene inserted in the AAVS1 region were obtained,where both Dsup mRNA and protein were expressed.The cell proliferation rate of HEK 293T-Dsup-copGFP was higher than that of HEK 293T-Control-copGFP(P＜0.001).Further investigation revealed that the expressions of Ki-67 and MCM4 protein in HEK 293T-Dsup-copGFP were significantly higher than in the control group,indicating that the knock in of Dsup gene might enhance the proliferation ability of human cells by promoting the expression of Ki-67 and MCM4 protein.Conclusion A gene editing vector is constructed,and stable cell line HEK 293T-Dsup-copGFP for Dsup fusion expression with copGFP is established.The expression of Dsup gene in HEK 293T cells can promote cell proliferation,possibly by upregulating the expressions of Ki-67 and MCM4 protein.

Background Air pollution is a major public health concern. Air Quality Health Index (AQHI) is a very important air quality risk communication tool. However, AQHI is usually constructed by single-pollutant model, which has obvious disadvantages. Objective To construct an AQHI based on the joint effects of multiple air pollutants (J-AQHI), and to provide a scientific tool for health risk warning and risk communication of air pollution. Methods Data on non-accidental deaths in Yunnan, Guangdong, Hunan, Zhejiang, and Jilin provinces from January 1, 2013 to December 31, 2018 were obtained from the corresponding provincial disease surveillance points systems (DSPS), including date of death, age, gender, and cause of death. Daily meteorological (temperature and relative humidity) and air pollution data (SO₂, NO₂, CO, PM_2.5, PM₁₀, and maximum 8 h O₃ concentrations) at the same period were respectively derived from China Meteorological Data Sharing Service System and National Urban Air Quality Real-time Publishing Platform. Lasso regression was first applied to select air pollutants, then a time-stratified case-crossover design was applied. Each case was matched to 3 or 4 control days which were selected on the same days of the week in the same calendar month. Then a distributed lag nonlinear model (DLNM) was used to estimate the exposure-response relationship between selected air pollutants and mortality, which was used to construct the AQHI. Finally, AQHI was classified into four levels according to the air pollutant guidance limit values from World Health Organization Global Air Quality Guidelines (AQG 2021), and the excess risks (ERs) were calculated to compare the AQHI based on single-pollutant model and the J-AQHI based on multi-pollutant model. Results PM_2.5, NO₂, SO₂, and O₃ were selected by Lasso regression to establish DLNM model. The ERs for an interquartile range (IQR) increase and 95% confidence intervals (CI) for PM_2.5, NO₂, SO₂ and O₃ were 0.71% (0.34%–1.09%), 2.46% (1.78%–3.15%), 1.25% (0.9%–1.6%), and 0.27% (−0.11%–0.65%) respectively. The distribution of J-AQHI was right-skewed, and it was divided into four levels, with ranges of 0-1 for low risk, 2-3 for moderate risk, 4-5 for high health risk, and ≥6 for severe risk, and the corresponding proportions were 11.25%, 64.61%, 19.33%, and 4.81%, respectively. The ER (95%CI) of mortality risk increased by 3.61% (2.93–4.29) for each IQR increase of the multi-pollutant based J-AQHI , while it was 3.39% (2.68–4.11) for the single-pollutant based AQHI . Conclusion The J-AQHI generated by multi-pollutant model demonstrates the actual exposure health risk of air pollution in the population and provides new ideas for further improvement of AQHI calculation methods.

OBJECTIVE: This study was aimed at investigating the carrier rate of, and molecular variation in, α- and β-globin gene mutations in Hunan Province. METHODS: We recruited 25,946 individuals attending premarital screening from 42 districts and counties in all 14 cities of Hunan Province. Hematological screening was performed, and molecular parameters were assessed. RESULTS: The overall carrier rate of thalassemia was 7.1%, including 4.83% for α-thalassemia, 2.15% for β-thalassemia, and 0.12% for both α- and β-thalassemia. The highest carrier rate of thalassemia was in Yongzhou (14.57%). The most abundant genotype of α-thalassemia and β-thalassemia was -α ^3.7/αα (50.23%) and β ^IVS-II-654/β ^N (28.23%), respectively. Four α-globin mutations [CD108 (ACC>AAC), CAP +29 (G>C), Hb Agrinio and Hb Cervantes] and six β-globin mutations [CAP +8 (C>T), IVS-II-848 (C>T), -56 (G>C), beta nt-77 (G>C), codon 20/21 (-TGGA) and Hb Knossos] had not previously been identified in China. Furthermore, this study provides the first report of the carrier rates of abnormal hemoglobin variants and α-globin triplication in Hunan Province, which were 0.49% and 1.99%, respectively. CONCLUSION Our study demonstrates the high complexity and diversity of thalassemia gene mutations in the Hunan population. The results should facilitate genetic counselling and the prevention of severe thalassemia in this region.
Humans ; beta-Thalassemia/genetics* ; alpha-Thalassemia/genetics* ; Hemoglobinopathies/genetics* ; China/epidemiology* ; High-Throughput Nucleotide Sequencing

Acute and chronic wounds seriously threaten patients' life health and quality of life, therefore, wound repair has become a hot topic of research for scholars at home and abroad in recent years. With the development of material science and tissue engineering, more and more biomaterials prepared from natural ingredients were used in basic research and clinical treatment of wound repair. Such biomaterials can be used as templates for wound tissue regeneration to induce autologous cell adhesion and migration, and promote the deposition of extracellular matrix, which have broad clinical application prospects. This paper reviews the characteristics and application advance of natural biomaterials which are popular in the field of wound repair, aiming to provide ideas for the research and development of new wound dressing and tissue engineering skin.

Objective: To investigate the clinical effects of ultra-pulsed fractional carbon dioxide laser (UFCL) in the treatment of mild to moderate microstomia after burns. Methods: A retrospective observational study was conducted on 19 patients with mild to moderate microstomia after burns who were admitted to Inner Mongolia Baogang Hospital from January 2018 to January 2022, including 15 males and 4 females aged (35±14) years. Patients had an average course of 71 d of microstomia, with 8 cases of moderate microstomia and 11 cases of mild microstomia. All the patients received UFCL treatment every 2-3 months until the microstomia was corrected or the treatment bottleneck was reached. The times of UFCL treatment for patients and the time interval from the last treatment to the last follow-up were recorded. Before the first treatment and at the last follow-up, the opening degree of mouth (finger measurement method), oral gap width, and the distance between the upper and lower incisors during mouth opening were recorded. Before the first treatment and at the last treatment, the new Vancouver scar scale (VSS) was used to evaluate the scar. At the last follow-up, the degree of satisfaction was evaluated by the Likert 5 scale by the patients themselves, and the satisfaction ratio was calculated; the adverse reactions such as pigmentation, blisters, infection, and persistent erythema in the treatment area were counted. Data were statistically analyzed with Mann-Whitney rank sum test or paired sample t test. Results: Patients received UFCL treatment of 3 (2, 6) times. The interval from the last treatment to the last follow-up was 26 months at most and 4 months at least. At the last follow-up, the opening degree of mouth of patients was significantly improved than that before treatment (Z=4.68, P<0.01). At the last follow-up, the oral gap width of patients was (35±6) mm, and the distance between upper and lower incisors during mouth opening was (3.2±0.4) cm, which was significantly improved compared with those before treatment (with t values of 10.73 and 18.97, respectively, P<0.01). The VSS score after the last treatment was 4.1±1.6, which was significantly better than that before treatment (t=22.96, P<0.01). At the last follow-up, the satisfaction ratio of patients with treatment was 18/19, and no pigmentation, blisters, infection, persistent erythema, and other adverse reactions of all patients in the treatment area occurred, however, one of the patients reported that the disease recurred about half a year after treatment. Conclusions: UFCL is an effective method for treating mild to moderate microstomia after burns, with which patients are highly satisfied, and it is worth of further study and promotion.
Blister ; Burns/therapy* ; Cicatrix/therapy* ; Female ; Humans ; Lasers, Gas/therapeutic use* ; Male ; Microstomia ; Treatment Outcome

With the aging of population and the development of social economy, the incidence of chronic wounds is increasing day by day, while the incidence of burns and trauma remains at a high level, making wound repair an increasingly concerned area in clinical practice. Thymosin β4 is a naturally occurring small molecule protein in vivo, which is widely distributed in a variety of body fluids and cells, especially in platelets. Thymosin β4 has biological activities of promoting angiogenesis, anti-inflammation, anti-apoptosis, and anti-fibrosis, and has many important functions in wound repair. Thymosin β4 has been observed to promote the healing of various wounds, such as burns, diabetic ulcers, pressure ulcers. This paper will review the molecular structure, mechanism of wound healing promotion, pharmacokinetics, and clinical application of thymosin β4, aiming to introduce its potential in wound treatment and the shortcomings of current researches.
Burns/drug therapy* ; Humans ; Pressure Ulcer ; Thymosin/therapeutic use* ; Wound Healing/physiology*

PURPOSE: This study was designed to compare the clinical efficacy of "8" and "0" wire fixation systems combined with double-head cannulated compression screws or Kirschner wires for the treatment of transverse patellar fractures. METHODS: From September 2011 to September 2018, patients with closed transverse patellar fractures treated with a double-head compression screw or Kirschner wire were included and analyzed retrospectively. Patients with patellar fractures combined with distal femoral fractures, tibial plateau fracture or preoperative lower limb dysfunction were excluded. The patients treated with the "8" tension band wire fixation system and Kirschner wire were taken as Group A; those treated with the "0" fixation system and Kirschner wire were taken as Group B; those treated with the "8" fixation system and double-head cannulated compression screw were taken as group C; and those treated with the "0" fixation system and double-head cannulated compression screw were taken as group D. Six weeks and one year after the operation and every month from the third month after the operation until the fractures healed, an X-ray examination was performed to identify fracture healing. The time of fracture healing and postoperative complications of the four groups were compared. One year after the operation, knee function was evaluated by Bostman's score. RESULTS: During the study period, 168 patients with patellar fractures were treated by operations, and 88 patients were excluded because the fracture type did not meet the requirements or because there were combined fractures of the distal femur or tibial plateau. As a result, 80 patients were included in this study, 20 in each group. All the patients were followed up for an average period of 12.2 months. Compared with Group A, patients in Group D presented less postoperative discomfort in the prepatellar region, quicker fracture healing, less fixation failure and better postoperative knee function scores (all p < 0.05). The incidence of internal fixation failure in Group (B+D) was lower than that in Group (A+C) (p > 0.05). CONCLUSION The "0" wire fixation system combined with a double-head cannulated compression screw seems to be more beneficial than the other three fixation systems for the treatment of transverse patellar fractures.

Objective:To explore the effect of cholesterol on the expression of genes for matrix synthesis and degradation of human meniscal fibrochondrocytes and its mechanism.Methods:Meniscal tissue was taken from patients undergoing arthroscopic surgery to extract fibrochondrocytes. The cells were divided into a control group in which the normal cells were not processed, a positive control group in which interleukin-1 β was used to create a degeneration model, and 2 treatment groups which were subjected to treatment with 15 and 30 μg/mL cholesterol respectively. Safranin O staining, β-galactosidase staining and enzymic kits were used to detect the morphology and total cholesterol (TCH) content of meniscal fibrochondrocytes in the 4 groups. Immunofluorescence and western blot were used to detect the protein expression of type Ⅰcollagen precursor α1 (COL1A1) and type Ⅱ collagen precursor α1 (COL2A1). RT-qPCR was used to detect the mRNA expression of COL1A1, COL2A1, matrix metalloproteinase (MMP) 3, MMP9, MMP13, and genes related to cholesterol efflux pathways [like liver X receptor α (LXR α), ATP binding cassette transporter A1 (ABCA1) and ABCG1]. Results:There was no significant difference between the control and the positive control groups in the TCH content in human meniscal fibrochondrocytes ( P>0.05). The treatments with 15 and 30 μg/mL cholesterol resulted in significantly increased TCH contents in human meniscal fibrochondrocytes in the treatment groups ( P<0.05). Compared with the control group, the mRNA expression of LXR α, ABCA1 and ABCG1 was significantly decreased in the treatment groups ( P<0.05), and the meniscal fibrochondrocytes in the positive group and the treatment groups presented with a lower density, chaotic distribution and obvious signs of degradation. Compared with the control groups, the mRNA expression of matrix synthesis genes (COL1A1 and COL2A1) in the meniscal fibrochondrocytes was significantly inhibited while the mRNA expression of matrix degradation metalloenzymes (MMP3, MMP9 and MMP13) was significantly promoted ( P<0.05). Conclusion:Cholesterol may inhibit the cholesterol efflux pathways of meniscal fibrochondrocytes, and thus cause accumulation of cholesterol in the meniscal fibrochondrocytes, eventually leading to degeneration of meniscus.

Objective:To explore the effects of Shenwei Ningyu pills （SNP）， a new Chinese medicine for depression， on the immunoinflammatory response mediated by Toll-like receptor 4 （TLR4）/myeloid differentiation factor 88 （MyD88） signaling pathway in the hippocampus of rats exposed to chronic restraint stress （CRS）. Method:Forty-four male Sprague Dawley rats were randomly enrolled into a normal group， a model group， an escitalopram group， and an SNP group. Except for the rats in the normal group， all rats were exposed to CRS and isolated rearing for 21 days continuously. Rats in the escitalopram group and the SNP group were administered with escitalopram （30 mg·kg^-1） and SNP （18 mg·kg^-1） one hour prior to CRS， respectively. The changes in body weight， sucrose preference index， horizontal movement scores， and vertical movement scores were observed by body weight assessment， sucrose preference test， and open field test. The expression of hippocampal TLR4 and MyD88 was detected by Western blot. The content of serum interleukin-1β （IL-1β）， IL-10， and tumor necrosis factor-α （TNF-α） was detected by enzyme-linked immunosorbent assay （ELISA）. Result:The results of the behavioral assessment showed that there was no significant difference in the changes of behavioral baselines among the groups before intervention. However， significant differences were found among the groups following different interventions. The body weight， sugar preference index， horizontal movement score， and vertical movement score of rats in the model group decreased after CRS for 21 days as compared with those in the normal group （P<0.01）. The above indicators in the SNP group and the escitalopram group were higher than those in the model group （P<0.01）， which indicated that SNP exerted an obvious antidepressant effect. The results of Western blot and ELISA showed that compared with the normal group， the model group showed elevated levels of hippocampal TLR4 and MyD88 and serum IL-1β and TNF-α （P˂0.01） and dwindled serum IL-10 （P˂0.01）， while SNP and escitalopram reversed the conditions in the model group （P˂0.01） except for TNF-α. Conclusion:The present study indicated that the antidepressant effect of SNP was presumedly achieved by inhibiting the immunoinflammatory response mediated by the TLR4/Myd88 signaling pathway in CRS rats.

Huangpu Tongqiao Capsules(HPTQC), with the functions of invigorating Qi and kidney, eliminating phlegm and removing blood stasis, have the effect of treating Alzheimer's disease(AD), but its mechanism needs further exploration. To explore the relationship between the therapeutic mechanism of HPTQC on Alzheimer's disease and EGFR-PLCγ signal pathway, 40 healthy male SD rats were selected and divided into 4 groups randomly: sham operation group(sham), model group(model), HPTQC group(HPTQC), and nimodipine group(NMP). AD rat model was established by intraperitoneal injection of D-galactose combined with an intracerebral injection of amyloid-β peptide(25-35). After 28 days of administration, Morris water maze test and HE staining showed that the learning and memory ability of AD rats were significantly decreased(P<0.01), and hippocampal neurons were obviously da-maged. However, HPTQC could improve the learning and memory ability of AD rats(P<0.05) and reduce the damage of hippocampal neurons. Immunofluorescence test results showed that the expression levels of EGFR and p-Tau in hippocampal CA1 region of AD rats were significantly increased(P<0.01), and HPTQC could reduce the expression of EGFR and p-Tau in hippocampus of AD rats(P<0.01). Western blot results showed that the protein expression levels of EGFR, PLCγ, IP3 R and p-Tau in hippocampus of AD rats were significantly increased(P<0.01), and HPTQC could reduce the protein expression of EGFR, PLCγ, IP3 R and p-Tau in AD rats(P<0.05). RT-PCR results showed that the mRNA levels of EGFR, PLCγ, IP3 R and Tau in hippocampus of AD rats were significantly increased(P<0.01), and HPTQC could reduce the mRNA levels of EGFR, PLCγ, IP3 R and Tau in AD rats(P<0.05). The results indicate that HPTQC can improve the learning and memory ability of AD rats, and its mechanism of action may be related to regulating EGFR-PLCγ signal pathway.
Alzheimer Disease ; Amyloid beta-Peptides ; Animals ; Capsules ; Disease Models, Animal ; ErbB Receptors ; Hippocampus ; Male ; Rats ; Rats, Sprague-Dawley ; Signal Transduction