Objective To observe the clinical effect of recombinant bovine basic fibroblast growth factor gel combined with esberitox for recurrent oral ulcer.Methods Forty-six patients with recurrent oral ulcer were divided into treatment group of 22 patients and control group of 24 patients by random digits table method.The patients in treatment group were given recombinant bovine basic fibroblast growth factor gel and esberitox.The patients in control group were given the watermelon frost spray only.After treatment for 1 week,the curative effect was compared.Results The cure rate in treatment group was 90.9％ (20/22),which was significantly higher than that in control group (66.7％,16/24),and there was significant difference (P ＜0.05).The time of pain vanish,oral ulcer decrease,oral ulcer healing in treatment group were significantly shorter than those in control group,and there were significant differences (P ＜0.05).Conclusions Recombinant bovine basic fibroblast growth factor gel combined with esberitox for recurrent oral ulcer can promote ulcer healing,shorten the course of disease,and has no adverse reaction.It is worthy of applying in clinic.

OBJECTIVE: To explore the role of N~6-methyladenosine(m~6A) catalytic enzymes(methyltransferases and demethylases) in cadmium-induced oxidative damage in human renal epithelial cells(HK-2 cells), and to analyze the correlation between nuclear factor-erythroid 2-related factor 2(NRF2) and m~6A catalytic enzymes. METHODS: i) HK-2 cells in logarithmic growth phase were randomly divided into control group and 6 cadmium sulfate treatment groups, then treated with 0, 2, 4, 8, 16, 32 and 64 μmol/L cadmium sulfate solution for 24 hours. The cell survival rates were detected by CCK-8 assay, and the appropriate doses of cadmium sulfate were selected for subsequent experiments. ii) HK-2 cells in logarithmic growth phase were randomly divided into control group and low-, medium-, and high-dose groups, and treated with 0, 4, 8, and 16 μmol/L cadmium sulfate solution respectively for 24 hours. Subsequently, the levels of reactive oxygen species(ROS) were detected by fluorescence probe. The mRNA expression of NRF2, the m~6A methyltransferases such as methyltransferase like proteins(METTL) 3, METTL14, METTL16 and the m~6A demethylases such as fat mass and obesity associated protein(FTO), AlkB family of nonheme Fe(Ⅱ)/α-ketoglutarate(α-KG)-dependent dioxygenases 5(ALKBH5) were determined by real-time polymerase chain reaction. RESULTS: i) The survival rate of HK-2 cells was more than 60.00% and lower than that of the control group(P<0.05) after the cells were stimulated with 16 μmol/L of cadmium sulfate. Therefore, 4, 8 and 16 μmol/L of cadmium sulfate were selected as the stimulation concentrations in the follow-up experiments. ii) The relative expression of NRF2, METTL3, METTL14 and METTL16 in HK-2 cells in low-dose group increased(all P<0.05), while the levels of ROS and the relative mRNA expression of NRF2, METTL3, METTL14, METTL16 and FTO in HK-2 cells in medium and high-dose groups increased(all P<0.05) when compared with the control group. There was no significant difference in the expression of ALKBH5 mRNA among these 4 groups(P>0.05). In the correlation analysis, NRF2 mRNA expression was positively correlated with the mRNA expression of METTL3 and METTL16 [Pearson correlation coefficient(r) = 0.61 and 0.66, respectively, all P<0.05]. There was no correlation between NRF2 mRNA expression and METTL14, FTO and ALKBH5(r=0.53, 0.48, and 0.01 respectively, all P>0.05). CONCLUSION: Cadmium sulfate may increase intracellular ROS level, up-regulate NRF2 expression and activate NRF2 signaling pathway as well as enhance the expression of METTL3 and METTL16 in HK-2 cells, thus increasing intracellular oxidative damage and decreasing the cell survival rate.

Acute Disease ; Female ; Gastrointestinal Hemorrhage ; etiology ; Humans ; Infant ; Intestine, Small ; abnormalities

Objective To analyze the relationship between the clinical and pathological effects and long-term prognosis in children with Henoch-Schonlein nephritis.Methods Changes of clinical pathology were studied in 32 children with Henoch-Schonlein nephritis and 19 cases of them were followed over an 8 to 14 year period.Results Acute nephritis ranked first(50 ?)and the nephritic syndrome ranked second(40?)in the clinical classification of Henoch-Schonlein nephritis;the majority had pathological changes of Grade???.The rate of recovery of acute nephritis and the nephritic syndrome was 55.6?abd 28.6?,respectively.The rate of recovery and deterioration of Grade??? pathological changes was 43.8?and 12.5?,respectively.Of the patients with Grade??? pathological changes,66.7? deteriorated or died. Conclusions The prognosis of acute nephritis was better than that of the nephritic syndrome,and long-term prognosis is closely associated with the clinical classification and pathology.

Objective To explore the association of cardiac disease associated genetic variants and the high incidence of Yunnan sudden unexplained death （YNSUD） in Yi nationality. Methods The genomic DNA was extracted from peripheral blood samples collected from 205 Yi villagers from YNSUD aggregative villages （inpatient group） and 197 healthy Yi villagers from neighboring villages （control group）. Fifty-two single nucleotide variants （SNVs） of 25 cardiac disease associated genes were genotyped using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS）. The SPSS 17.0 was used to analyze data. The pathogenicities of variants with differences between the two groups that have statistical significance were predicted by protein function prediction software PolyPhen-2 and SIFT. All villagers from inpatient group were given electrocardiogram （ECG） examination using a 12-lead electrocardiograph. Results The allele frequency and the genotype frequency of missense mutation DSG2 （rs2278792, c.2318G>A, p.R773K） of pathogenic genes of arrhythmogenic right ventricular cardiomyopathy （ARVC） in inpatient group was higher than that in control group （P<0.05）. Abnormal ECG changes were detected in 71 individuals （34.6%） in the inpatient group, among which 54 individuals carried R773K mutation, including clockwise （counterclockwise） rotation, left （right） axis deviation, ST segment and T wave alteration and heart-blocking. Conclusion Definite pathogenic mutations have not been found in the 52 cardiac disease genes associated SNVs detected in Yi nationality in regions with high incidence of YNSUD. The cause of high incidence of YNSUD in Yi nationality needs further study.
Arrhythmogenic Right Ventricular Dysplasia ; China/epidemiology* ; Death, Sudden/etiology* ; Death, Sudden, Cardiac/etiology* ; Ethnicity/genetics* ; Humans ; Incidence ; Mutation

Objective:The aim of this study is to compare the effects of age on the biological properties of adipose-derived stem cells(ASCs) and fat survival of ASC-assisted lipotransfer. To identify the effect of age factors on the biological characteristics of human ASCs and compare the effects of ASCs-assisted subcutaneous fat transplantation/lipotransfer on nude mice at different ages.Methods:Human lipoaspirates were obtained from 30 healthy female patients (aged from 18 to 65 years) acquiring the abdominal liposuction. Samples were divided into three groups according to donor age: group A, 18-29 years; group B, 30-49 years; group C, 50-65 years. Stromal vascular fraction cells were isolated from the harvested adipose tissue using collagenase. The yield and cell viability of SVF were tested using the Muse cell count and viability assay. ASCs were cultured and harvested at the second passage. MSC surface markers of ASCs were examined by the flow cytometry. The cell proliferation of ASCs from different age was determined by the CCK-8 assay. The scratch test was used for assessing the ASCs migration ability. The adipogenic differentiation potential of ASCs was analyzed by induction of lipid formation in vitro. The expression levels of PPAR-γ and CEBP-α genes were detected by RT-PCR assay. The survival of adipocytes in the grafts was analyzed by perilipin-A immunofluorescence staining. The fat survival of ASCs-enriched grafts from different age was measured in animal models. The weight and residual volume of fat grafts were compared in different groups after three months. The histologic analysis was evaluated by cell integrity and necrosis tissue in fat grafts. The vessel density was measured using the CD31 immunohistochemical staining. Data were analyzed by SPSS software version 21.0 with one-way ANOVA to compare the difference of multiple groups. A value of P< 0.05 was considered statistically significant. Results:The yield and cell viability of SVF isolated from lipoaspirates were: group A, (7.06±1.28)×10 5/ml and 82.46%±2.81%; group B, (6.90±0.32)×10 5/ml and 82.01%±3.85%; group C, (6.40±0.62)×10 5/ml and 77.82%±3.45%, respectively. No significant difference was found in different age groups. SVF viability was decreased with increasing age. The expression of positive surface markers CD90, CD44, CD105 and CD73 of ASCs in each group was above 95%, and the expression of negative surface markers was below 2%, all of which met the criteria for the expression level of mesenchymal stem cell surface markers. Moreover, there was a decline in cell proliferation and migration of ASCs with increasing age. No significant difference was found in the adipogenic differentiation of ASCs in three groups. The fat grafts were harvested three months after cell-assisted lipotransfer. The graft weight was(0.18±0.02) g in group A, (0.17±0.02) g in group B, (0.15±0.01) g in group C, (0.13±0.03) g in control group, respectively; F=9.274, P<0.001. The residual volume of grafts was(262.88±17.69)/mm 3 in group A, (263.83±25.96)/mm 3 in group B, (240.06±25.08)/mm 3 in group C, (201.81±31.48)/mm 3 in the control group; F=12.95, P<0.001. There were significant differences in the weight and residual volume of fat grafts in different age groups( F=5.231, P=0.012; F=3.364, P=0.049). HE staining result showed that compared with the blank control group, ASC-assisted groups had uniform distribution of adipocytes, less fibrous connective tissue and necrotic tissue. There was a statistically significant difference in the proportion of fat integrity and necrotic tissue between the groups ( F=3.434, P=0.027; F=9.314, P<0.001). Results of the histologic analysis showed no significant difference in the proportion of fat cell integrity and necrotic tissue in each group( F=0.282, P=0.756; F=0.421, P=0.661). Immunofluorescence staining result showed that, compared with the control group, a higher number of perilipin-positive adipocytes were observed in ASCs-assisted fat grafting from different age groups, with uniform distribution. The vessel density of fat grafts was (15.70±4.16)/mm 2 in group A, (17.03±8.30)/mm 2 in group B; (16.68±6.71)/mm 2 in group C, (11.50±4.04)/mm 2 in control group; F=3.523, P=0.019. Conclusions:The proliferation and migration of human ASCs decreased with age, but age did not affect the adipogenic differentiation potential of ASCs. ASCs from different ages effectively improved the fat survival of grafts. ASCs-assisted fat grafting was more effective in young people than in elder.

Objective:The aim of this study is to compare the effects of age on the biological properties of adipose-derived stem cells(ASCs) and fat survival of ASC-assisted lipotransfer. To identify the effect of age factors on the biological characteristics of human ASCs and compare the effects of ASCs-assisted subcutaneous fat transplantation/lipotransfer on nude mice at different ages.Methods:Human lipoaspirates were obtained from 30 healthy female patients (aged from 18 to 65 years) acquiring the abdominal liposuction. Samples were divided into three groups according to donor age: group A, 18-29 years; group B, 30-49 years; group C, 50-65 years. Stromal vascular fraction cells were isolated from the harvested adipose tissue using collagenase. The yield and cell viability of SVF were tested using the Muse cell count and viability assay. ASCs were cultured and harvested at the second passage. MSC surface markers of ASCs were examined by the flow cytometry. The cell proliferation of ASCs from different age was determined by the CCK-8 assay. The scratch test was used for assessing the ASCs migration ability. The adipogenic differentiation potential of ASCs was analyzed by induction of lipid formation in vitro. The expression levels of PPAR-γ and CEBP-α genes were detected by RT-PCR assay. The survival of adipocytes in the grafts was analyzed by perilipin-A immunofluorescence staining. The fat survival of ASCs-enriched grafts from different age was measured in animal models. The weight and residual volume of fat grafts were compared in different groups after three months. The histologic analysis was evaluated by cell integrity and necrosis tissue in fat grafts. The vessel density was measured using the CD31 immunohistochemical staining. Data were analyzed by SPSS software version 21.0 with one-way ANOVA to compare the difference of multiple groups. A value of P< 0.05 was considered statistically significant. Results:The yield and cell viability of SVF isolated from lipoaspirates were: group A, (7.06±1.28)×10 5/ml and 82.46%±2.81%; group B, (6.90±0.32)×10 5/ml and 82.01%±3.85%; group C, (6.40±0.62)×10 5/ml and 77.82%±3.45%, respectively. No significant difference was found in different age groups. SVF viability was decreased with increasing age. The expression of positive surface markers CD90, CD44, CD105 and CD73 of ASCs in each group was above 95%, and the expression of negative surface markers was below 2%, all of which met the criteria for the expression level of mesenchymal stem cell surface markers. Moreover, there was a decline in cell proliferation and migration of ASCs with increasing age. No significant difference was found in the adipogenic differentiation of ASCs in three groups. The fat grafts were harvested three months after cell-assisted lipotransfer. The graft weight was(0.18±0.02) g in group A, (0.17±0.02) g in group B, (0.15±0.01) g in group C, (0.13±0.03) g in control group, respectively; F=9.274, P<0.001. The residual volume of grafts was(262.88±17.69)/mm 3 in group A, (263.83±25.96)/mm 3 in group B, (240.06±25.08)/mm 3 in group C, (201.81±31.48)/mm 3 in the control group; F=12.95, P<0.001. There were significant differences in the weight and residual volume of fat grafts in different age groups( F=5.231, P=0.012; F=3.364, P=0.049). HE staining result showed that compared with the blank control group, ASC-assisted groups had uniform distribution of adipocytes, less fibrous connective tissue and necrotic tissue. There was a statistically significant difference in the proportion of fat integrity and necrotic tissue between the groups ( F=3.434, P=0.027; F=9.314, P<0.001). Results of the histologic analysis showed no significant difference in the proportion of fat cell integrity and necrotic tissue in each group( F=0.282, P=0.756; F=0.421, P=0.661). Immunofluorescence staining result showed that, compared with the control group, a higher number of perilipin-positive adipocytes were observed in ASCs-assisted fat grafting from different age groups, with uniform distribution. The vessel density of fat grafts was (15.70±4.16)/mm 2 in group A, (17.03±8.30)/mm 2 in group B; (16.68±6.71)/mm 2 in group C, (11.50±4.04)/mm 2 in control group; F=3.523, P=0.019. Conclusions:The proliferation and migration of human ASCs decreased with age, but age did not affect the adipogenic differentiation potential of ASCs. ASCs from different ages effectively improved the fat survival of grafts. ASCs-assisted fat grafting was more effective in young people than in elder.

OBJECTIVES: To explore the cytotoxicity of four wild mushrooms involved in a case of Yunnan sudden unexplained death (YNSUD), to provide the experimental basis for prevention and treatment of YNSUD. METHODS: Four kinds of wild mushrooms that were eaten by family members in this YNSUD incident were collected and identified by expert identification and gene sequencing. Raw extracts from four wild mushrooms were extracted by ultrasonic extraction to intervene HEK293 cells, and the mushrooms with obvious cytotoxicity were screened by Cell Counting Kit-8 (CCK-8). The selected wild mushrooms were prepared into three kinds of extracts, which were raw, boiled, and boiled followed by enzymolysis. HEK293 cells were intervened with these three extracts at different concentrations. The cytotoxicity was detected by CCK-8 combined with lactate dehydrogenase (LDH) Assay Kit, and the morphological changes of HEK293 cells were observed under an inverted phase contrast microscope. RESULTS: Species identification indicated that the four wild mushrooms were Butyriboletus roseoflavus, Boletus edulis, Russula virescens and Amanita manginiana. Cytotoxicity was found only in Amanita manginiana. The raw extracts showed cytotoxicity at the mass concentration of 0.1 mg/mL, while the boiled extracts and the boiled followed by enzymolysis extracts showed obvious cytotoxicity at the mass concentration of 0.4 mg/mL and 0.7 mg/mL, respectively. In addition to the obvious decrease in the number of HEK293 cells, the number of synapses increased and the refraction of HEK293 cells was poor after the intervention of Amanita manginiana extracts. CONCLUSIONS The extracts of Amanita manginiana involved in this YNSUD case has obvious cytotoxicity, and some of its toxicity can be reduced by boiled and enzymolysis, but cannot be completely detoxicated. Therefore, the consumption of Amanita manginiana is potentially dangerous, and it may be one of the causes of the YNSUD.
Humans ; HEK293 Cells ; Sincalide ; China ; Amanita ; Death, Sudden

Light adaptation enables the vertebrate visual system to operate over a wide range of ambient illumination. Regulation of phototransduction in photoreceptors is considered a major mechanism underlying light adaptation. However, various types of neurons and glial cells exist in the retina, and whether and how all retinal cells interact to adapt to light/dark conditions at the cellular and molecular levels requires systematic investigation. Therefore, we utilized single-cell RNA sequencing to dissect retinal cell-type-specific transcriptomes during light/dark adaptation in mice. The results demonstrated that, in addition to photoreceptors, other retinal cell types also showed dynamic molecular changes and specifically enriched signaling pathways under light/dark adaptation. Importantly, Müller glial cells (MGs) were identified as hub cells for intercellular interactions, displaying complex cell‒cell communication with other retinal cells. Furthermore, light increased the transcription of the deiodinase Dio2 in MGs, which converted thyroxine (T4) to active triiodothyronine (T3). Subsequently, light increased T3 levels and regulated mitochondrial respiration in retinal cells in response to light conditions. As cones specifically express the thyroid hormone receptor Thrb, they responded to the increase in T3 by adjusting light responsiveness. Loss of the expression of Dio2 specifically in MGs decreased the light responsive ability of cones. These results suggest that retinal cells display global transcriptional changes under light/dark adaptation and that MGs coordinate intercellular communication during light/dark adaptation via thyroid hormone signaling.
Animals ; Mice ; Dark Adaptation ; Light ; Retina ; Retinal Cone Photoreceptor Cells/metabolism* ; Adaptation, Ocular ; Neuroglia/physiology* ; Cell Communication ; Thyroid Hormones