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RESUMEN La acondroplasia severa con retraso del desarrollo y acantosis nigricans (SADDAN) es una rara y letal displasia esquelética. Presentamos el primer caso detectado en Perú, en un infante de 13 meses con características fenotípicas de macrocefalia relativa, tórax estrecho, extremidades micromélicas y piel en acordeón; asimismo, un marcado retraso del desarrollo psicomotor en todos los hitos (prueba peruana) y acantosis nigricans. El paciente tuvo mala evolución clínica caracterizada por crisis convulsivas recurrentes, dificultad respiratoria progresiva, y falleció por insuficiencia respiratoria concomitante a neumonía. Esta entidad requiere del acceso a exámenes específicos como el panel de displasias esqueléticas, la cual no es parte de la oferta en la mayoría de los hospitales del Perú. Se requiere una mayor atención las enfermedades raras, a fin de proveer diagnósticos e información oportuna a los involucrados.
ABSTRACT Severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN) is a rare and lethal skeletal dysplasia. We present the first case detected in Peru, in a 13-month-old infant with phenotypic characteristics of relative macrocephaly, narrow thorax, micromelic extremities and accordion skin; likewise, a marked delay in psychomotor development in all milestones (Peruvian test), and acanthosis nigricans. The patient had a poor clinical evolution characterized by recurrent seizures, progressive respiratory difficulty, dying from respiratory failure concomitant to pneumonia. This entity requires access to specific exams such as the skeletal dysplasia panel, which is not part of the offering in most hospitals in Peru. Greater attention is required for rare diseases, to provide timely diagnoses and information to those involved.
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Objective To explore the expression of junctophilin 2(JP2)and fibroblast growth factor 23(FGF23)in a rabbit model of atrial fibrillation mediated-cardiomyopathy(AMC).Methods Rabbit models of atrial fibrillation(AF)were developed through rapid atrial stimulation and then divided into three groups:control group(pacemak-ers implanted without pacing,n=6),AF group(pacing with ejection fraction decrease＜10％,n=5),and AMC group(pacing with ejection fraction decrease≥10％,n=6).Echocardiography was performed to detect left ventric-ular end-diastolic diameter(LVEDD),left ventricular end-systolic diameter(LVESD)and left ventricular ejection fraction(LVEF).JP2 and FGF23 were detected by ELISA method.Western blot and RT-qPCR were conducted to detect protein and mRNA expression of JP2 and FGF23.Results Left atrial diameter,right atrial diameter and right ventricular diameter increased and LVEF decreased in the AMC group as compared with the control group.AMC group had lower LVEF and larger aorta and right ventricle diameter.Compared with the control group,the ex-pression of FGF23(P＜0.001)and JP2(P＜0.01)in left atrial cardiomyocytes was significantly increased in the AF group,while the expression of JP2 was decreased in the AMC group(P＜0.001).AMC group had lower expression of JP2 and FGF23 compared with AF group.Compared to the control group,plasma concentration of JP2 and FGF23 increased in the AF group and FGF23 plasma concentration increased in the AMC group.Plasma concentration of FGF23 and JP2 was lower in AMC group than that in AF group.Conclusions FGF23 expression increased and JP2 expression decreased as found in the rabbit AMC model.
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Objective·To evaluate the changes in cognitive function in overweight and obese adolescents,and explore the association between cognitive function and fibroblast growth factor 21(FGF21).Methods·A total of 175 adolescents from a senior high school in Shanghai were divided into normal weight group(n=50),overweight group(n=50)and obese group(n=75)based on their body mass index(BMI).General information,anthropometric data and laboratory testing indicators of the adolescents were collected and compared.The cognitive function of the three groups of adolescents was assessed by using the accuracy(ACC)and reaction time of Flanker task and n-back task.Enzyme-linked immunosorbent assay(ELISA)was used to detect the serum FGF21 level of the three groups of adolescents.Partial correlation analysis and multiple linear regression model were used to evaluate the correlation between cognitive task performance and anthropometric data and laboratory testing indicators.Results·Compared with the normal weight group,systolic blood pressure,diastolic blood pressure,and the levels of fasting plasma glucose,glycosylated hemoglobin and triacylglycerol in the obese group were higher(all P<0.05).Under congruent or incongruent stimulus conditions in the Flanker task,there was no significant difference in ACC between any two groups;compared with the normal weight and overweight groups,the reaction time of the adolescents in the obese group was prolonged(all P<0.05).In the n-back task,there were no significant differences in ACC between any two groups,while the obese group had longer reaction time in the 1-back and 2-back tasks compared to the normal weight and overweight groups(all P<0.05).Compared with the normal weight group,serum FGF21 levels of the adolescents in the obese group were higher(P=0.000).Partial correlation analysis showed that the reaction time of the adolescents in Flanker and n-back tasks was correlated with their BMI,body fat mass,waist circumference,waist-to-hip ratio and FGF21 level(all P<0.05).Multiple linear regression analysis further confirmed that BMI was associated with prolonged reaction time in cognitive-related behavioral tasks in the adolescents(all P<0.05),and FGF21 level was associated with ACC in the 2-back task(P=0.000)and reaction time in the incongruent stimulus condition(P=0.048).Conclusion·Overweight and obese adolescents have cognitive impairments,and BMI and serum FGF21 levels are associated with changes in their cognitive function.
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BACKGROUND:At present,effective preventive and therapeutic measures for hypertrophic scar are still limited.In contrast,most of botanical herbs have few side effects and abundant sources,offering new ideas and approaches for the prevention and treatment for hypertrophic scar. OBJECTIVE:To explore the potential molecular mechanism of plant-derived β-sitosterol on hypertrophic scar fibroblasts by network pharmacology and molecular docking techniques and to initially verify it by cytological experiments. METHODS:Through the network pharmacology,the relevant database and software were used to screen the drug targets of β-sitosterol and obtain the hypertrophic scar-related disease targets.The potential(intersection)targets of β-sitosterol on hypertrophic scar were obtained.Cytoscape software and STRING database were used to construct the"drug-target-disease"network and protein-protein interaction network,and screen out the core targets in the protein-protein interaction network.Gene ontology(GO)biological function and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment analyses of intersection targets were conducted through the DAVID database,and the signaling pathways and core target genes closely related to the intersection targets were further identified through literature analysis.AutoDock software was used to perform the molecular docking of β-sitosterol and core target proteins.In vitro cellular assays were used to verify the effects of β-sitosterol on proliferation,apoptosis,cell cycle distribution and mRNA expression of core target genes in human hypertrophic scar fibroblasts. RESULTS AND CONCLUSION:There were 56 intersection targets of β-sitosterol and hypertrophic scar and 10 core targets were identified in the protein-protein interaction network,including tyrosine kinase,mitogen-activated protein kinase 3(MAPK3),cysteine protease 3(CASP3),apolipoprotein E,estrogen receptor 1,sterol regulatory element-binding transcription factor 1,peroxisome proliferator-activated receptor alpha,C-reactive protein,intercellular adhesion molecule 1,and catalase.Combined with the literatures and the functional analysis of the KEGG and GO,the MAPK signaling pathway was further identified to be closely related to the intersection targets,and MAPK3(ERK1-MAPK),CASP3,P53 and tumor necrosis factor were identified as the core targets.The molecular docking results indicated that β-sitosterol was well bound to the core target proteins.Cellular assays showed that 100 μmol/L β-sitosterol inhibited hypertrophic scar fibroblast proliferation,decreased mitochondrial membrane potential and induced apoptosis(P<0.01),increased the proportion of G1-phase cells and decreased the proportion of S-phase cells(P<0.05),upregulated the mRNA expression of CASP3,P53 and tumor necrosis factor(P<0.05),and downregulated the mRNA expression of MAPK3(P<0.001).To conclude,β-sitosterol may induce cell apoptosis in hypertrophic scar fibroblasts by activating the tumor necrosis factor pathway and upregulating the expression of CASP3 and P53,while inhibiting the ERK-MAPK pathway to arrest cell cycle and thus reduce the proliferation of hypertrophic scar fibroblasts.
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BACKGROUND:Abnormal extracellular matrix accumulation and excessive proliferation of fibroblasts are the main manifestations of pathological scars.Excessive proliferation of fibroblasts leads to the production of large amounts of collagen-based extracellular matrix.Therefore,to investigate the role of fibroblast fibrosis in the formation of pathological scar will provide a new idea for revealing the mechanism of pathological scar and biological therapy. OBJECTIVE:To investigate the effect of RAS-selective lethal small molecule 3(RSL3)on the fibrosis of human pathological scar fibroblasts. METHODS:Then cases of pathological scar tissue and normal skin tissue samples from the same individuals,provided by the Department of Burn Plastic Surgery,General Hospital of Ningxia Medical University,were collected.Fibroblasts of human pathological scar and human normal skin were extracted and used in the following experiments.The general condition of the pathological scar tissue and the normal skin tissue was detected by hematoxylin-eosin staining.The appearance of fibroblasts from pathological scar and normal skin were observed by inverted microscope.The fibroblasts were verified by immunofluorescence assay.The cells were treated with different concentrations of RSL3(1,3,5,7,9,11,13 μmol/L).The inhibitory concentration of RSL3 on fibroblasts was detected by cell counting kit-8.Control group(without treatment)and RSL3 intervention group(treated with 7 μmol/L RSL3 for 24 hours)were set up.The mRNA and protein expressions of glutathione peroxidase 4,type Ⅰ collagen,type Ⅲ collagen and α-smooth muscle actin were detected by Qrt-PCR and western blot,respectively.Level of malondialdehyde in cells was detected.The residual scratch area was measured by cell scratch test after 24 hours to calculate the percentage of residual scratch area. RESULTS AND CONCLUSION:The expression of glutathione peroxidase 4 in the pathological scar group was higher than that in the normal skin group(Mrna:t=3.252,P<0.01;protein:t=5.075,P<0.01).The expression of glutathione peroxidase 4 in the pathological scar fibroblast group was higher than that in the normal skin fibroblast group(Mrna:t=10.32,P<0.01;protein:t=26.22,P<0.01).Compared with the control group,the expression of glutathione peroxidase 4 was decreased(Mrna:t=2.798,P<0.05;protein:t=4.643,P<0.01),the content of malondialdehyde was increased(t=2.917,P<0.05),the expression of type Ⅰ collagen(Mrna:t=15.84,P<0.01;protein:t=4.610,P<0.01),type Ⅲ collagen(Mrna:t=28.86,P<0.01;protein:t=7.713,P<0.01)and α-smooth muscle actin(Mrna:t=2.671,P<0.05;protein:t=7.417,P<0.01)were decreased in the RSL3 intervention group.Compared with the control group,the migration ability was weakened in the RSL3 intervention group(t=14.06,P<0.01).To conclude,RSL3 can inhibit the expression of glutathione peroxidase 4 and then inhibit the ability of fibrosis and migration of pathological scar fibroblasts.
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BACKGROUND:Mangiferin is a biphenylpyridone compound extracted from mango leaves,bark and roots.Previous studies have shown that mangiferin can exert anti-systemic inflammatory effects through the activation of transcription factors such as NF-κB and JAK/STAT. OBJECTIVE:To investigate the effects and mechanisms of mangiferin on proliferation,migration and inflammatory factor release of rheumatoid arthritis fibroblast-like synovial cells(RA-FLS). METHODS:RA-FLS were divided into blank group,R848(TLR7/8 agonists)stimulated group,mangiferin low-,medium-,high-dose groups(2,4 and 8 μg/mL)and positive control group(Cu-CPT8,TLR8 pathway inhibitor).The cytotoxic effect of different mass concentrations of mangiferin was detected using cell counting kit-8 method and the final cellular dosing mass concentration was screened.The proliferation ability of RA-FLS was detected by cell clone formation assay,the migration ability of RA-FLS was detected by scratch assay and Transwell migration assay,and the expression of interleukin 1β,interleukin 6 and tumor necrosis factor α mRNA in RA-FLS was detected by qRT-PCR. RESULTS AND CONCLUSION:Compared with the blank group,the viability of RA-FLS was inhibited after treatment with mangiferin at 2-10 μg/mL,but there was no significant difference among groups(P>0.05),indicating that the toxic effect on RA-FLS was minimal.Compared with the R848-stimulated group,mangiferin decreased the number of cell clones,the scratch healing rate and the number of migrating cells in all dosing groups(P<0.01);and the expression of interleukin 1β,interleukin 6 and tumor necrosis factor α mRNA was also reduced in the mangostin medium-and high-dose groups(P<0.01).Compared with the R848-stimulated group,the number of cell clones,the scratch healing rate and the number of migrating cells as well as the expression levels of interleukin 6 and tumor necrosis factor α mRNA were significantly reduced in the positive control group(P<0.05,P<0.01).But there was no significant difference in the expression level of interleukin 1β.To conclude,mangiferin may exert its anti-rheumatoid arthritis effects through the TLR7/8 signaling pathway by inhibiting RA-FLS proliferation,migration,and inflammatory factor release.
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BACKGROUND:Hypertrophic scar is a skin fibrosis disease characterized by excessive proliferation of fibroblasts,epidermal thickening,and stratum corneum dysfunction.At present,the pathogenesis of Hypertrophic scar is still unclear. OBJECTIVE:To screen the core(Hub)genes and important signaling pathways in hypertrophic scar-related datasets based on bioinformatics,and then verify them by cell experiments to predict small molecule drugs that may have therapeutic effects on hypertrophic scar. METHODS:Datasets related to hypertrophic scar were searched from Gene Expression Omnibus(GEO)database,and differentially expressed genes were identified by R software analysis.Gene ontology and KEGG enrichment analyses were performed for differentially expressed genes.Protein-protein interaction network of differentially expressed genes was constructed using String online platform.Then,the key genes and core modules in the protein-protein interaction network were screened by Cytohubba and MCODE plugin-in Cytoscape software respectively,and the Hub genes were obtained by the intersection of the above key genes and the genes that formed the core module.Real-time fluorescent quantitative PCR was used to verify the difference in Hub gene mRNA expression between human hypertrophic scar and normal skin epidermal stem cells.The histological data from the Human Protein Atlas were used to verify the differences in the expression and distribution of Hub gene-encoded proteins in the two kinds of human tissues.Finally,the potential drugs for hypertrophic scar were predicted by the connectivity map database. RESULTS AND CONCLUSION:Among the identified differentially expressed genes,102 genes were up-regulated and 702 genes were down-regulated.Gene ontology and KEGG analysis showed that the enriched signaling pathways and biological processes were mainly involved in tight junction,arachidonic acid metabolism,extracellular matrix receptor interaction,epidermal development and keratinization.Eight Hub genes were found to be closely related to the mevalonate pathway that regulates cholesterol metabolism,including HMGCS1,DHCR7,MSMO1,FDPS,MVK,HMGCR,MVD and ACAT2.Compared with the normal skin group,the mRNA expression of HMGCS1,DHCR7,MSMO1,FDPS,HMGCR,MVD and ACAT2 in the hypertrophic scar group decreased significantly(P＜0.05),while MVK mRNA expression had no significant change(P＞0.05).Except for MVK,the expression levels of other Hub gene-encoded proteins in normal skin tissue were higher than those in hypertrophic scar tissue(P＜0.05).The top 10 candidate drugs included protein kinase A inhibitor(H-89),serine protease inhibitor(Dabigatran-Etexilate),FLT3 inhibitor(sunitinib),among which resveratrol and β-sitosterol are plant extracts.To conclude,Hub genes closely related to mevalonate metabolism may affect the structure and function of the epidermis by regulating lipid metabolism,which may an important pathogenesis of hypertrophic scar.The small-molecule compounds identified in this study can be used as candidate drugs for the treatment of hypertrophic scar.
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BACKGROUND:There are many studies focusing on keloid scars,but the pathogenesis is not fully understood.In recent years,there have been some new research advances in the pathogenesis of keloids,including transforming growth factor-β(TGF-β)/Smad signaling pathway,ischemic hypoxia,hypoxia-inducible factor 1(HIF-1),and mitogen-activated protein kinase(MAPK)pathway.The TGF-β/Smad pathway is now more clearly studied,and activation of the TGF-β/Smad pathway promotes the development of keloid scars. OBJECTIVE:To review the TGF-β/Smad signaling pathway and evaluate the main therapeutic strategies targeting this pathway,with the aim of contributing to the development of more effective clinical treatments. METHODS:PubMed and Web of Science,CNKI and WanFang databases were searched by computer for relevant literature published from January 2017 to April 2023 with the search terms of"keloid,fibroblasts,TGF-β/Smad,extracellular matrix,collagen,treatment measures"in English and Chinese.Seventy-two articles were finally included according to the inclusion and exclusion criteria. RESULTS AND CONCLUSION:The mechanism of TGF-β/Smad signaling pathway in the occurrence and development of keloids is summarized:TGF-β1 and TGF-β2 are overexpressed in keloids,while TGF-β3 shows antifibrotic effects.Smad2/3 and Smad1/5/8 are combined with Smad4 to form a complex that enters the nucleus and plays a fibrotic role,while Smad6/7 can inhibit keloid hyperplasia.The TGF-β/Smad signaling pathway is currently the most clearly studied pathway in keloids,and there are many pathways targeted to inhibit the activation of this pathway,which can inhibit the occurrence and development of keloids to a greater extent.Currently,there is no single clinical gold standard treatment for keloids,and inhibition of the TGF-β/Smad pathway alone cannot completely inhibit the development of keloids.A comprehensive consideration of the association between all systemic systems and keloids is needed.Although many promising targets have been identified in the fibrosis cascade,more research is needed to translate this into targeted therapies in the clinic.
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BACKGROUND:Previous studies have shown that salidroside has an ameliorative effect on multi-organ fibrosis.However,the protective effect of salidroside on angiotensin ⅱ-induced fibrosis in cardiac fibroblasts is unclear. OBJECTIVE:To investigate the protective effects of salidroside on angiotensin ⅱ-induced oxidative stress and extracellular matrix deposition in cardiac fibroblasts of Sprague-Dawley rats and its mechanism of action. METHODS:Angiotensin Ⅱ was used to induce fibrosis in cardiac fibroblasts,and there were five experimental groups:normal control group,model group(final concentration of angiotensin Ⅱ in culture medium was 1 μmol/L),salidroside low and high dose groups(treatment with salidroside 50,100 μmol/L for 2 hours,followed by co-incubation with angiotensin Ⅱ for 48 hours),SIRT1 inhibitor group(treatment with SIRT1 inhibitor EX527 10 μmol/L for 2 hours,followed by high dose of salidroside for 2 hours and then co-incubation with angiotensin Ⅱ for 48 hours).The cell viability was detected using the cell counting kit-8 method,the cell migration rate was detected by Transwell,the intracellular reactive oxygen species level was detected by DCFH-DA fluorescent probe,and the intracellular malondialdehyde content,superoxide dismutase and catalase activities were detected by relevant kits.The protein and mRNA expression levels of SIRT1,LOXL2,α-SMA,type I collagen and type Ⅲ collagen were detected by western blot and qRT-PCR,respectively. RESULTS AND CONCLUSION:The cells were identified as cardiac fibroblasts by Vimentin fluorescence.Compared with the normal control group,cell viability,cell migration rate,reactive oxygen species level,and malondialdehyde content were significantly increased,superoxide dismutase and catalase activities were significantly decreased,LOXL2,α-SMA,type I collagen,type Ⅲ collagen mRNA and protein expression were significantly increased,and SIRT1 protein expression level was significantly decreased in the model group(all P＜0.01).Compared with the model group,the above indexes showed opposite changes in the salidroside low and high dose groups(all P＜0.05).Moreover,salidroside showed dose-dependent regulation.Compared with salidroside groups,cell migration rate and α-SMA protein expression level were significantly increased in the SIRT1 inhibitor group(both P＜0.001).To conclude,salidroside has a protective effect on angiotensin Ⅱ-induced cardiac fibroblasts and can dose-dependently inhibit oxidative stress and extracellular matrix deposition.
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BACKGROUND:Combining seed cells with 3D bioprinting technology enables the specific construction of various tissues and organs to meet the demands of tissue repair.However,further research is needed on the promotion of angiogenesis in damaged tissues. OBJECTIVE:By cultivating a 3D scaffold structure of methacrylated gelatin loaded with fibroblasts,obtaining the supernatant,and mixing it in different proportions with a complete culture medium to simulate the cellular microenvironment during tissue repair,this study aimed to explore the role of various cellular microenvironments in promoting angiogenesis in endothelial cells. METHODS:A methacrylated gelatin scaffold structure loaded with fibroblasts was prepared using an extrusion-based 3D bioprinting process.Hydrogel scaffold extract was prepared and mixed with a complete culture medium in ratios of 1:1,1:2,and 1:4 to obtain conditioned medium.Mouse embryonic fibroblasts BALB3T3 and human umbilical vein endothelial cells were co-cultured with complete medium(control group)and hydrogel scaffold extract,respectively.Cell proliferation was assessed using the CCK-8 assay and cell viability was analyzed using live/dead staining.Three kinds of conditioned medium and complete medium(control group)were used to co-culture with human umbilical vein endothelial cells for tube formulation assay,vascular genetic testing,and immunofluorescence staining of CD31. RESULTS AND CONCLUSION:(1)Scanning electron microscopy revealed that the methacrylated gelatin scaffold exhibited a porous structure,and rheological results demonstrated excellent mechanical properties of the hydrogel.CCK-8 assay and live/dead cell staining showed that the hydrogel scaffold extract had no obvious cytotoxicity.(2)Tube formulation assay indicated that the hydrogel showed the total length of cell tubules in 1:1 conditioned medium group was smaller than that in the control group(P<0.05).There were no statistical differences among the four groups in the number of vascular branches formed by endothelial cells(P>0.05).(3)qRT-PCR results showed that for vascular endothelial growth factor mRNA expression,the 1:2 conditioned medium group was lower than the 1:1 conditioned medium group on day 1(P<0.01).On day 3,the expression level of vascular endothelial growth factor in the 1:2 conditioned medium group was higher than that in the control group(P<0.01).On day 5,the cytokine expression level in the 1:2 conditioned medium group was significantly higher than that in the other three groups(P<0.01 or P<0.000 1).The expression in the 1:1 conditioned medium group was significantly lower than that in the other three groups(P<0.05 or P<0.01).On day 1,the expression level of basic fibroblast growth factor in the 1:1 conditioned medium group was significantly higher than that in the control group and 1:4 conditioned medium group(P<0.01,P<0.05).The expression was higher in the 1:2 conditioned medium group than that in the control group(P<0.05).On day 3,the expression levels of cytokines in the 1:4 conditioned medium group was higher than that in the control group(P<0.05).(4)On day 3,the expression of CD31 in the 1:2 conditioned medium group was higher than that in the control group and the 1:4 conditioned medium group(P<0.05).(5)The results indicate that the resulting conditioned media can simulate the microenvironment of vascular regeneration after tissue damage,promoting the vascularization process of endothelial cells.The best promotion of vascularization in endothelial cells was observed when the ratio of supernatant to complete culture medium was 1:2.
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BACKGROUND:The temporal and spatial expression of fibroblast growth factor receptors 1 and 2 remains a controversial issue during kidney development,so the relationship between them and kidney development remains unclear. OBJECTIVE:To observe the dynamic expression of fibroblast growth factor receptors 1 and 2 during kidney development of mice,and to investigate the relationship between them and kidney development. METHODS:The kidneys of fetal mice[embryotic days(E)12,14,16,and 18]and neonatal mice[neonatal days(N)1,3,7,14,24,and 40]were selected to examine the temporal and spatial expression of fibroblast growth factor receptors 1 and 2 by immunohistochemistry method in kidney tissues,and quantitative analysis was performed using western blot assay. RESULTS AND CONCLUSION:(1)Immunohistochemistry showed that fibroblast growth factor receptor 1 was mainly localized in metanephric tissue surrounding the tip of the ureteral bud at E12.Subsequently,fibroblast growth factor receptor 1 was expressed in immature renal corpuscles at various stages,some distal convoluted tubules and capillary loops.The positive site was mainly concentrated in the generative region.Fibroblast growth factor receptor 2 was initially expressed in both ureteral buds and metanephric tissue.Fibroblast growth factor receptor 2 was localized in immature renal corpuscles,distal tubules,collecting ducts and thin segments of medullary loops with kidney development.However,the expression of renal corpuscles was weak.(2)Stereology and western blot assay showed that the expression of fibroblast growth factor receptor 1 was high before birth and gradually decreased after birth,while the expression was very low after N7 day.The expression level of fibroblast growth factor receptor 2 increased gradually with the kidney development and tended to be stable after N7 day.(3)The results exhibit that fibroblast growth factor receptors 1 and 2 are expressed spatially and temporally during kidney development.It is speculated that fibroblast growth factor receptors 1 and 2 may influence nephron development and maturation,and fibroblast growth factor receptor 2 is critical during the formation of ureteral buds and morphology.
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BACKGROUND:Heterotopic ossification of skeletal muscle is a clinically serious complication.For heterotopic ossification of skeletal muscles,the cells involved in the process of heterotopic ossification remain unclear. OBJECTIVE:To investigate the involvement of myocytes,fascia cells,and endothelial cells in the process of heterotopic ossification in skeletal muscle and to observe the cell origin of heterotopic ossification in skeletal muscle induced by bone morphogenetic protein 4. METHODS:Both C2C12 cells and the myotubes formed by the C2C12 cells in the induction medium were cultured,and 500 ng/mL bone morphogenetic protein 4 was added to the medium respectively,and whether the C2C12 cells and myotubes continued to proliferate within 10 days under the treatment were observed under a microscope.Myogenic cells(L6,derived from rats)and fibroblast-derived cells(derived from human)were co-cultured.After treatment with 500 ng/mL bone morphogenetic protein 4 and 10 ng/mL transforming growth factor-β,osteogenic and chondrogenic differentiation potential within 21 days were observed using Safranine O staining and Alcian blue staining.Using transgenic animal FVB/N-TgN(TIE2-LacZ)182Sato mice,15 μL of adeno-associated virus-bone morphogenetic protein 4(5×1010 PFU/mL)were implanted in the thigh muscle space of genetic mice for 10 and 14 days.X-gal staining was used to observe the formation of new blood vessel endothelium in the differentiated bone. RESULTS AND CONCLUSION:(1)Bone morphogenetic protein 4 caused myotube breakdown and increased C2C12 cell proliferation.Compared with other groups,the pure fibroblast-derived cell group had a higher area of positive alcian blue and safarin O staining(P<0.05)and a lower area of alkaline phosphatase staining(P<0.05),while the pure L6 group had a bigger area of alkaline phosphatase staining(P<0.05)but a smaller area of positive alcian blue and safarin O staining(P<0.05).(2)Transplantation of adeno-associated virus-bone morphogenetic protein 4-adsorbed gelatin sponge into FVB/N-TgN(TIE2-LacZ)182Sato mice resulted in heterotopic ossification.(3)X-gal staining results demonstrated that there was no obvious staining in chondrocytes and differentiated bones and Tie2+ endothelial cells did not participate in the formation of the alienated bone.(4)These findings verify that fibroblasts are the primary source of osteoblasts during the adeno-associated virus-bone morphogenetic protein 4-induced ectopic endochondral ossification in skeletal muscle,but myogenic cells are the main source of osteoblasts.Tie2+ endothelial cells might not be the cell source for cartilage and bone.
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BACKGROUND:Heterotopic ossification is a dynamic growth process.Diverse heterotopic ossification subtypes have diverse etiologies or induction factors,but they exhibit a similar clinical process in the intermediate and later phases of the disease.Acquired heterotopic ossification produced by trauma and other circumstances has a high incidence. OBJECTIVE:To summarize the molecular biological mechanisms linked to the occurrence and progression of acquired heterotopic ossification in recent years. METHODS:The keywords"molecular biology,heterotopic ossification,mechanisms"were searched in CNKI,Wanfang,PubMed,Embase,Web of Science,and Google Scholar databases for articles published from January 2016 to August 2022.Supplementary searches were conducted based on the obtained articles.After the collected literature was screened,131 articles were finally included and summarized. RESULTS AND CONCLUSION:(1)The occurrence and development of acquired heterotopic ossification is a dynamic process with certain concealment,making diagnosis and treatment of the disease difficult.(2)By reviewing relevant literature,it was found that acquired heterotopic ossification involves signaling pathways such as bone morphogenetic protein,transforming growth factor-β,Hedgehog,Wnt,and mTOR,as well as core factors such as Runx-2,vascular endothelial growth factor,hypoxia-inducing factor,fibroblast growth factor,and Sox9.The core mechanism may be the interaction between different signaling pathways,affecting the body's osteoblast precursor cells,osteoblast microenvironment,and related cytokines,thereby affecting the body's bone metabolism and leading to the occurrence of acquired heterotopic ossification.(3)In the future,it is possible to take the heterotopic ossification-related single-cell osteogenic homeostasis as the research direction,take the osteoblast precursor cells-osteogenic microenvironment-signaling pathways and cytokines as the research elements,explore the characteristics of each element under different temporal and spatial conditions,compare the similarities and differences of the osteogenic homeostasis of different types and individuals,observe the regulatory mechanism of the molecular signaling network of heterotopic ossification from a holistic perspective.It is beneficial to the exploration of new methods for the future clinical prevention and treatment of heterotopic ossification.(4)Meanwhile,the treatment methods represented by traditional Chinese medicine and targeted therapy have become research hotspots in recent years.How to link traditional Chinese medicine with the osteogenic homeostasis in the body and combine it with targeted therapy is also one of the future research directions.(5)At present,the research on acquired heterotopic ossification is still limited to basic experimental research and the clinical prevention and treatment methods still have defects such as uncertain efficacy and obvious side effects.The safety and effectiveness of relevant targeted prevention and treatment drugs in clinical application still need to be verified.Future research should focus on clinical prevention and treatment based on basic experimental research combined with the mechanism of occurrence and development.
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BACKGROUND:Mesenchymal stem cells are susceptible to senescence during in vitro expansion,which greatly hinders their application in vivo and in vitro.How to improve the replicative senescence of mesenchymal stem cells is an urgent problem to be solved in tissue engineering. OBJECTIVE:To determine whether vascular endothelial growth factor combined with basic fibroblast growth factor can improve the aging of bone marrow mesenchymal stem cells caused by replicative passage. METHODS:Rat bone marrow mesenchymal stem cells were extracted by whole bone marrow adhesion method.Passage 2 cells were selected as normal control group.Passage 7 and later algebraic cells were selected as aging model group.Vascular endothelial growth factor(50 μg/L),basic fibroblast growth factor(10 μg/L),and their combination were administered.Cell proliferation was detected by CCK-8 assay.Cell senescence was observed by β-galactosidase activity staining.Cytoskeleton size and colony formation ability were observed by phalloidine staining and Giemsa staining,respectively,and the levels of senescence-related genes P16,P21,and P53 were detected by qRT-PCR.Gene expression levels of P16,P21,and P53 were tested by qRT-PCR. RESULTS AND CONCLUSION:(1)Vascular endothelial growth factor combined with basic fibroblast growth factor could promote the proliferation of aged bone marrow mesenchymal stem cells,which began to enter the plateau stage on day 9,and the absorbance value of the combined intervention group was significantly higher than that of the model group on day 9(P<0.05).(2)The phenotypic markers of the cells in the combined intervention group did not change,and the cell morphology changed from broad to slender.(3)Compared with the model group,the positive rate of β-galactosidase was significantly decreased(P<0.01);the number of nuclei increased(P<0.001);the total area of cytoskeleton increased(P<0.01);colony formation ability was enhanced(P<0.05);expression level of P16 was decreased(P<0.01)in the combined intervention group.These results indicate that vascular endothelial growth factor combined with basic fibroblast growth factor can improve the senescence of bone marrow mesenchymal stem cells caused by replicative passage without changing the cell phenotype.
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Objective To investigate the expression level,diagnostic value and correlation of miR-497-5p and human fibroblast growth factor-2(FGF-2)in patients with Alzheimer's disease(AD).Methods The clinical data of 50 patients with first diagnosed AD and 37 normal subjects(control group)were collected,among which AD patients were divided into mild AD group(n=18),moder-ate AD group(n=18)and severe AD group(n=14).The expression level of miR-497-5p was detected by real-time quantitative polymerase chain reaction(RT-qPCR)and FGF-2 was detected by enzyme-linked immunosorbent assay(ELISA).Mini-mental state examination(MMSE)was used to evaluate the cognitive function of AD patients,and the correlation between miR-497-5p and MMSE and FGF-2 levels was analyzed.The diagnostic efficacy of miR-497-5p and FGF-2 levels for AD was evaluated using receiv-er operator characteristic(ROC)curve.Results Compared with the control group and mild AD group,the expression levels of miR-497-5p in moderate and severe AD groups were significantly increased(P＜0.01),and the level of FGF-2 was significantly decreased(P＜0.01).MiR-497-5p in AD group was negatively correlated with MMSE score and FGF-2 level(r were-0.724 and-0.748,P＜0.01).ROC curve analysis results showed that miR-497-5p,FGF-2 and their combined indexes had higher area under the curve,sensitivity and specificity in the diagnosis of moderate and severe AD and in the differentiation of mild and moderate AD,as well as mild and severe AD,and the combined indexes of miR-497-5p and FGF-2had the best diagnostic and differential efficacy.Conclusion Serum miR-497-5p is up-regulated and FGF-2 level is down-regulated in patients with moderate and severe AD.The combined detection of miR-497-5P and FGF-2has certain diagnostic value for moderate and severe AD and provides certain reference.
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AIM:To observe how total flavonoids of Pterocarya hupehensis Skan(PHSTF)affects the migra-tion and invasion of human rheumatoid fibroblast-like synoviocyte line MH7A.METHODS:The MH7A cells were divided into control group(without any treatment),low-,medium-and high-dose(6.25,12.5 and 25 mg/L,respectively)PHSTF groups,phosphatidylinositol 3-kinase(PI3K)inhibitor 740Y-P(10 μmol/L)group,and 740Y-P(10 μmol/L)+high-dose(25 mg/L)PHSTF group.The viability of the MH7A cells was determined by CCK-8 assay,while the migration and inva-sion were assessed by scratch and Transwell assays.The protein levels of matrix metalloproteinase-2(MMP-2),MMP-9,PI3K,p-PI3K,AKT and p-AKT were detected by Western blot.RESULTS:Compared with control group,the cell via-bility in PHSTF treatment groups was reduced(P<0.05),the cell wound healing area was significantly decreased(P<0.01),migratory and invasive cells in the lower chamber were significantly reduced(P<0.01),and the protein expres-sion of MMP-2 and MMP-9 and the ratios of p-PI3K/PI3K and pAKT/AKT were decreased(P<0.01).Compared with high-dose PHSTF group,the addition of PI3K/AKT pathway agonist 740Y-P significantly increased the migration and invasion ability of MH7A cells(P<0.01)and elevated the protein expression of MMP-2 and MMP-9 and the ratios of p-PI3K/PI3K and pAKT/AKT(P<0.01)under the treatment with PHSTF.CONCLUSION:Total flavonoids of Pterocarya hupehensis Skan could inhibit the migration and invasion of MH7A cells by regulating the PI3K/AKT signaling pathway.
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Objective:To investigate the efficacy of pulp revascularization in the treatment of pulp necrosis with periapical periodontitis in young permanent teeth and its effect on the levels of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in gingival crevicular fluid.Methods:From January 2021 to August 2021, 72 young patients with permanent teeth exhibiting pulp necrosis and apical periodontitis who were treated at Haiyang People's Hospital were included in this study. These patients were subsequently divided into a study group ( n = 35) and a control group ( n = 37), depending on their respective treatment methods. The control group underwent conventional apical angioplasty, whereas the study group underwent pulp revascularization. A comparative analysis was conducted to assess the clinical efficacy of both treatments. Additionally, levels of VEGF and bFGF in gingival crevicular fluid were measured before and after surgery, and these values were compared between the two groups. Relevant clinical indicators and the incidence of adverse reactions were also compared between the study and control groups. Results:The overall response rate in the study group was 94.3% (33/35), which was significantly higher than 70.3% (26/37) in the control group ( χ2 = 7.01, P < 0.05). Prior to surgery, there were no notable differences in VEGF level, bFGF level, root length, or root canal thickness between the two groups (all P > 0.05). However, after surgery, VEGF level, bFGF level, root length, and root canal thickness in the study group were (43.25 ± 4.87) ng/L, (40.72 ± 4.83) ng/L, (8.95 ± 0.27) mm, and (3.08 ± 0.24) mm, respectively. These values were (39.90 ± 4.80) ng/L, (36.05 ± 4.66) ng/L, (8.55 ± 0.18) mm, and (2.90 ± 0.20) mm, respectively, in the control group. There were significant differences in VEGF level, bFGF level, root length, and root canal thickness between the two groups ( t = 2.96, 4.18, 5.67, 2.88, all P < 0.05). After surgery, the scores for apical inflammation, root development, and Visual Analogue Scale (VAS) in the study group were significantly higher than those in the control group ( t = 7.61, 4.83, 9.47, all P < 0.001). The incidence of adverse reactions in the study group was 2.9% (1/35), which was significantly lower than 21.6% (8/37) in the control group ( χ2 = 5.79, P < 0.05). Conclusion:Pulp revascularization exhibits superior curative effects compared with conventional apical angioplasty for the treatment of pulp necrosis and apical periodontitis in young permanent teeth. This treatment effectively alleviates pain, markedly improves tooth function, and has a low incidence of adverse reactions, highlighting its clinical value as a therapeutic option.
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Objective:To investigate the efficacy of combined repair therapy using recombinant bovine basic fibroblast growth factor (R-bFGF) gel and silver ion dressing on the donor site of patients with hand trauma undergoing skin grafting.Methods:Eighty patients with hand trauma who underwent skin grafting at Lishui Central Hospital between October 2020 and October 2021 were enrolled in this study. Using a simple random grouping method, the patients were randomly assigned to a control group and an observation group in a 1:1 ratio, with 40 patients in each group. The control group received conventional vaseline gauze treatment, while the observation group was treated with a combination of R-bFGF gel and silver ion dressing. After 2 weeks of treatment, the repair effects of both groups were evaluated. Before and after treatment, the Connor-Davidson Resilience Scale (CD-RISC) scores and Visual Analog Scale (VAS) scores were compared between the control and observation groups. Additionally, wound healing time, granulation tissue growth time, wound epithelium formation time, and dressing change times as well as total active motion of the fingers were evaluated and compared between the two groups.Results:The repair effect in the observation group was significantly superior to that in the control group ( Z = 4.92, P < 0.05). Furthermore, the recovery of hand function in the observation group was notably better than that in the control group ( Z = 4.31, P < 0.05). The CD-RISC score in the observation group was significantly higher than that in the control group [(77.54 ± 11.35) points vs. (70.61 ± 9.72) points, t = 2.93, P < 0.05]. Additionally, the VAS score, wound healing time, granulation tissue growth time, wound epithelium formation time, and dressing change times in the observation group were significantly lower or fewer than those in the control group [(4.95 ± 1.13) points vs. (5.52 ± 1.24) points, (10.43 ± 1.65) days vs. (15.54 ± 1.71) days, (7.42 ± 2.35) days vs. (11.56 ± 2.71) days, (10.25 ± 2.47) days vs. (12.82 ± 2.64) days, and (2.12 ± 0.63) times vs. (3.35 ± 0.86) times, t = -2.15, -13.60, -7.30, -4.50, -7.30, all P < 0.05]. Conclusion:The combined use of R-bFGF gel and silver ion dressing effectively enhances the repair outcomes of skin donor sites, thereby improving the psychological well-being and reducing pain perception in patients with hand trauma. This therapeutic approach markedly promotes the prognosis and functional recovery of these patients, offering valuable clinical reference for the treatment of hand injuries.
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Objective:To explore the effect of low intensity pulsed ultrasound(LIPUS)on inhibiting the abnormal cell phenotype of fibroblast-like synoviocytes in rheumatoid arthritis(RA-FLS)and possible mechanism. Method:Synoviocytes were isolated by using enzyme digestion,and the morphology of cells was observed un-der microscope.At the same time,the expression of Vimentin protein was detected by immunofluorescence method to identify RA-FLS.Cells cultured in vitro were divided into four groups:control group,LIPUS group,tumor necrosis factor(TNF-α)group and TNF-α+LIPUS group or three groups:control group,interleu-kin-6(IL-6)group and IL-6+LIPUS group.The effects of LIPUS on RA-FLS cell viability and proliferation were detected by CCK8 and EDU assay respectively,and the effects of LIPUS on RA-FLS migration were ob-served by scratch test and Transwell migration assay.RT-qPCR was used to detect the gene expression of im-portant cytokines,chemokines and matrix metalloproteinases(MMPs)in RA-FLS.ELISA was used to further detect the effect of LIPUS on the expression of IL-6,a key effector of RA-FLS,and the effects of LIPUS on mitogen-activated protein kinase(MAPK)signaling pathway in RA-FLS were detected by Western Blot. Result:Purified RA-FLS were obtained.Firstly,LIPUS could suppress the cell activity(P＜0.00l)and prolifer-ation(P=0.007)induced by TNF-α in RA-FLS cultured in vitro.However,the migration and the transcription levels of MMPs related to migration(MMP2 and MMP9)were not significantly different between groups(P＞0.05).LIPUS could inhibit the high expression of IL-6 and interleukin-8(IL-8)at the mRNA level in the in-flammatory environment induced by TNF-α(P＜0.001),but there was no significant difference in the suppres-sion of interleukin-1β(IL-1β),MMP1 and MMP13(P＞0.05).In addition,compared with untreated group,LI-PUS could inhibit the secretion of IL-6 in RA-FLS induced by TNF-α(P＜0.001),and also inhibited the pro-liferation of RA-FLS induced by IL-6(P-0.003).Finally,LIPUS could inhibit the phosphorylation of p38 MAPK and c-Jun N-terminal kinase(JNK)in MAPK signaling pathway(P=0.033),but the effect on the phosphorylation of extracellular signal-regulated kinas 1/2(ERK1/2)was not significantly(P＞0.05). Conclusion:LIPUS could reduce the abnormal proliferation of RA-FLS in inflammatory state without affecting its migration,which might be related to the inhibition of p38/JNK-IL-6 signaling pathway.
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OBJECTIVE Based on fibroblast activation protein(FAP)gene promoter as the response element,to develop a new dual luciferase reporting system for the screening of drugs related to myocardial fibrosis.METHODS The promoter fragment of mouse FAP gene was synthesized in vitro and cloned into plasmid psiCHECK2 to replace HSV-TK promoter,and then a new recombinant plasmid psiCHECK2-FAP was obtained.After the recombinant plasmid psiCHECK2-FAP was digested by restriction endonucliase Hind Ⅲ,the product digested was identified by agar-gel electrophoresis and sequencing.After psiCHECK2-FAP was transient transfected into mouse cardiac fibroblasts(MCFs),and continued cultured for 24 h,and MCFs were treated with Ransforming growth factor β1(TGF-β1,5 μg·L-1)or angiotensinⅡ(Ang Ⅱ,1 μmol·L-1)or palmitic acid(PA,100 μmol·L-1)for 0,12,24,48 h,respectively,the double luciferase reporter gene assay was used to detect luciferase activity;After psiCHECK2-FAP was transient transfected into MCFs,the cells were pretreated with Dapa(1 μmol·L-1)for 1 h,and supplemented with TGF-β1(5 μg·L-1)or AngⅡ(1 μmol·L-1)or PA(100 μmol·L-1),continued treatment for 24 h,the double luciferase reporter gene assay was used to detect luciferase activity,and the expression of collagenⅠand collagen Ⅲ protein was detected with Western blotting.RESULTS The recombinant plasmid psiCHECK2-FAP was digested into two fragments by Hind Ⅲ with the expected strip size,and the sequencing results were completely consistent with the theoretical sequence;Compared with control group,the collagenⅠand collagen Ⅲ protein expression were significantly increased by TGF-β1 or Ang Ⅱ or PA in MCFs(P<0.05,P<0.01).However,compared with TGF-β1 or Ang Ⅱ or PA group,the intervention of Dapa significantly alleviated the promoter activity of FAP gene and the expression of collagenⅠand collagen Ⅲ protein(P<0.05,P<0.01);Compared with control group,luciferase activity was significantly increased by TGF-β1 or Ang Ⅱ or PA(P<0.05,P<0.01),reaching the peak at 24 h.Compared with TGF-β1 or AngⅡ or PA group,the intervention of Dapa significantly decreased luciferase activity(P<0.05,P<0.01).CONCLUSSION Based on the promoter of FAP gene as the response element,a noval dual luciferase reporter gene system was established and showed a good sensitivity to the promyocardial fibrosis factor in MCFs,which can provide strategies for the development of antimyocar-dial fibrosis drugs.