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@#Abstract: The differential expression and subcellular localization of single-cell proteins are closely related to the physiological state and pathological mechanisms of the body. The development of single-cell protein in situ imaging methods provides powerful tools for spatial single-cell proteomics research and single-cell protein profiling. This article summarizes the single-cell protein imaging methods developed in recent years, including the circulating immunofluorescence imaging methods based on ordered multi-round antibody incubation, mass spectrometry imaging based on metal element labeled antibodies, fluorescence imaging based on DNA-barcoded antibody, gene encoded fluorescence protein imaging and spectral imaging based on Raman spectroscopy or X-ray spectroscopy, with brief explanation of the imaging principles of these methods. It focuses on the multiple performance, imaging resolution and signal amplification performance of these methods, and analyzes their application characteristics in practical scientific research and clinical work, in the hope of providing some reference for the development of more revolutionary single-cell imaging methods, and promoting the development of biomedical and precision medicine.
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Objective To explore the clinical significance of nucleolar antinuclear antibodies(ANA)in re-lated diseases.Methods This study was a retrospective study.Clinical samples of 71780 patients who visited the hospital from January 2017 to May 2022 were collected.Indirect immunofluorescence was used to detect ANA in clinical samples.Statistical analysis was conducted on the positivity rate of nucleolar ANA in clinical patients,as well as the relevant clinical information and laboratory characteristics of patients with autoimmune diseases(AID)with nucleolar ANA positivity.Results Among 71780 patients who underwent routine ANA testing,16778 were positive for ANA,with a positive rate of 23.37％.Among them,there were 1 708 cases of nucleolar type,accounting for 2.38％of all routine ANA tests,and the proportion of ANA positive cases was 10.18％.There was a statistically significant difference in the positive rate of nucleolar ANA between patients of different genders in the＞20-＜50 year old group and the ≥ 50 year old group(P＜0.05),while there was no statistically significant difference in the positive rate of nucleolar ANA between patients of different genders in the ≤ 20 year old group(P＞0.05).There was a statistically significant difference in the positivity rate of nucleolar ANA among women of different age groups(P＜0.05),among them,the highest positive rate of nucleolar ANA was found in women aged between 20 and 50 years old.There was no statistically significant difference in the positive rate of nucleolar ANA among males of different age groups(P＞0.05).The positivi-ty rate of ANA was the highest among patients in the Department of Rheumatology and Immunology(70.35％),but nucleolar ANA positivity was mainly seen in departments such as Reproductive Medicine Cen-ter(12.90％),Respiratory Medicine(12.40％),and Neurology(11.29％),and the difference in positivity rates between departments was statistically significant(P＜0.05).Out of 1 708 nucleolar ANA positive indi-viduals,420 underwent ANA titers,including 34 AID patients and 386 non AID patients.There was no statis-tically significant difference in nucleolus positive titers between non AID patients and AID patients(P＞0.05).Conclusion The nucleolus type is a common fluorescence pattern in ANA positive individuals,and there are gender and age differences in ANA positive individuals.The positive rate and titer of nucleolar ANA vary among different AID diseases.Combined with other immune function indicators,and it is helpful for early differential diagnosis of AID.
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Objective To investigate the effects of neuroligin-1,-2(NLGN-1,-2)on oligodendrocyte(OLs)differentiation and myelination in the central nervous system.Methods OLs were cultured in vitro in the presence of different concentrations of NLGN-1 and NLGN-2.Morphological differentiation of OLs was observed by immunofluorescent staining and mRNA expression levels of myelin-associated genes were detected by Real-time PCR.Western blotting was used to detect the expression of myelin-related proteins.Results NLGN-1,-2 accelerated the differentiation of oligodendrocyte precursor cells(OPCs)into mature OLs,and promoted the ability of myelin sheath formation.In vitro culture conditions,the dosage of 500 μg/L had the best promotion effect on OLs differentiation and maturation,and NLGN-2 had better promoting effect than that of NLGN-1.Furthermore,the mRNA expression levels of myelin-associated genes myelin protein P0(MPZ),myelin basic protein(MBP)increased after the neuroligins treatments detected by Real-time PCR.Western blotting result showed that the expressions of MBP and MPZ increased significantly after 500 μg/L treatment with NLGN-1 and NLGN-2 for 12 hours.Conclusion NLGN-1,-2 promote OLs differentiation and myelination.The positive effect of NLGN-2 is greater than that of NLGN-1 significantly,suggesting that the treatment with inhibitory synaptic-associated cytokines may improve the ability of myelin sheath formation in the central nervous system.
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Objective To explore the effect of scutellarin on lipopolysaccharide(LPS)induced neuroinflammation in BV-2 microglia cells.Methods BV-2 microglia were cultured and randomly divided into 6 groups:control group(Ctrl),cyclic GMP-AMP synthetase(cGAS)inhibitor RU320521 group(RU.521 group),LPS group,LPS+RU.521 group,LPS+scutellarin pretreatment group(LPS+S)and LPS+S+RU.521 group.The expressions of cGAS,stimulator of interferon gene(STING),nuclear factor kappa B(NF-κB),phosphorylated NF-κB(p-NF-κB),neuroinflammatory factors PYD domains-containing protein 3(NLRP3)and tumor necrosis factor α(TNF-α)in BV-2 microglia were detected by Western blotting and immunofluorescent double staining(n= 3).Results Western blotting and immunofluorescent double staining showed that compared with the control group,the expression of cGAS,STING,p-NF-κB,NLRP3 and TNF-α in BV-2 microglia increased significantly after LPS induction(P<0.05),while the expression of cGAS,STING,p-NF-κB,NLRP3 and TNF-α in LPS+S group were significantly lower than those in LPS group(P<0.05).Treatment with cGAS pathway inhibitor RU.521 showed similar effects as the pre-treatment group with scutellarin.In addition,the change of NF-κB in each group was not statistically significant(P>0.05).Conclusion Scutellarin inhibits the neuroinflammation mediated by BV-2 microglia cells,which may be related to cGAS-STING signaling pathway.
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Objective To discuss the relationship between activated glia cells in distal segment of the spinal cord and widespread pain.Methods Fifty female rats were randomly divided into sham group,the chronic constriction injury of the infraorbital nerve(CCI-ION)group,CCI-ION+minocycline(Mino)group,CCI-ION+L-2-aminoadipic acid(LAA)group,and CCI-ION+normal saline(NS)group,n=10 for each group.CCI-ION model was established and Mino,LAA,and normal saline were delivered intrathecally to CCI-ION rats.Immunofluorescence staining was used to detect activated astrocytes and microglia in the medulla oblongata,cervical,thoracic,and lumbar spinal cord segments.On the 7th,14th,21st,28th day,von Frey filaments were used to evaluate the mechanical withdrawal threshold of vibrissa pad,and electronic von Frey tactile pain meter was used to measure the mechanical withdrawal threshold of front paw,chest and hind paw.The radiant thermal stimulator was used to measure the thermal withdrawal threshold of hind paw.Results After intrathecal injection of Mino to inhibit microglia,the activated microglia in each spinal cord segment decreased.Moreover,inhibiting astrocytes by using LAA significantly reduced activated astrocytes in spinal dorsal horn from distal segments.Behavioral assay showed that after intrathecal injection of Mino and LAA,the mechanical allodynia of vibrissa pad in CCI-ION rats was relieved.However,there was no significant difference(P>0.05)in the thermal and mechanical withdrawal thresholds in the hind paw of CCI-ION rats after intrathecal injection of Mino,while intrathecal injection of LAA significantly increased the thermal and mechanical withdrawal thresholds in the hind paw,indicating the relief of widespread pain induced by CCI-ION.Conclusion The activated astrocytes in distal segments of the spinal cord mediated CCI-ION-induced widespread pain.
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Objective To investigate the effect of glucagon-like peptide 1(GLP-1)receptor agonists exendin-4 on the secretion of cyclophilin A(CyPA)to inhibit atherosclerosis(AS)and vascular calcification in mice role of the process.Methods Twenty ApoE-/-mice were randomly divided into model group and exendin-4 group,10 mice in each group,and were fed with high-fat diet to establish AS model,another 10 wild-type C57BL/6J mice were taken as the control group,and the exendin-4 group was intraperitoneally injected with the GLP-1R agonist exendin-4,1/d,for 8 weeks.After 8 weeks,the ELISA method was used to determine the level of triglyceride(TG),total cholesterol(TC),high density lipoprotein cholesterol(HDL-C),low density lipoprotein cholesterol(LDL-C)and CyPA,serum calcium level was detected by methylthymol blue colorimetric method,oil red O staining to detect the development of atherosclerotic plaques in the aorta,HE staining was used to observe the pathological changes of the aorta,Von Kossa staining was used to observe the calcium deposition in the aorta,immunohistochemical staining,Real-time PCR and Western blotting were used to detect the expression levels of aortic RUNX2 and bone morphogenetic protein 2(BMP-2),immunofluorescent staining was used to detect the positive expression of CyPA in aortic tissue.Results Compared with the control group,the serum levels of TG,TC,LDL-C,Ca and CyPA in the model group increased(P<0.05),the atherosclerotic plaque areas of the aorta increased(P<0.05),the aortic wall was thickened significantly and a large number of inflammatory cells were infiltrated,a large amount of calcium deposits were deposited in the aortic parietal membrane,the positive expression area ratio of RUNX2 and BMP-2,the relative mRNA expression of RUNX2 and BMP-2,the relative protein expression of RUNX2 and BMP-2 in aortic tissue all increased(P<0.05),and the red fluorescence of CyPA expression in aortic tissue was enhanced significantly.Compared with the model group,the serum levels of TG,TC,LDL-C,Ca and CyPA in the exendin-4 group decreased(P<0.05),the atherosclerotic plaque areas of the aorta decreased(P<0.05),the thickening of the aortic wall and the infiltration of inflammatory cells were alleviated significantly,the calcium deposition in the aortic wall was reduced,the positive expression area ratio of RUNX2 and BMP-2,the relative mRNA expression of RUNX2 and BMP-2,the relative protein expression of RUNX2 and BMP-2 in aortic tissue all decreased(P<0.05),and at the same time,the red fluorescence of CyPA expression in aortic tissue was weakened significantly.Conclusion GLP-1 receptor agonists exendin-4 can inhibit atherosclerosis and vascular calcification in mice,and the mechanism may be related to the reduction of CyPA secretion.
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Objective To prepare and characterize the mouse polyclonal antibody against the dense granule protein 24 (GRA24) of Toxoplasma gondii, and explore its preliminary applications. Methods The GRA24 coding sequences of different T. gondii strains were aligned using the MEGA-X software, and the dominant peptide of the GRA24 protein was analyzed with the Protean software. The base sequence encoding this peptide was amplified using PCR assay and ligated into the pET-28a vector, and the generated GRA24 truncated protein was transformed into Escherichia coli BL21. After induction by isopropyl-beta-D-thiogalactopyranoside (IPTG), the expression and purification of the recombinant GRA24 protein was analyzed using sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE). BALB/c mice were immunized by subcutaneous injection with the purified recombinant GRA24 truncated protein to generate the polyclonal antibody, and the titer of the polyclonal antibody was measured using enzyme linked immunosorbent assay (ELISA). The specificity of the polyclonal antibody was tested using Western blotting, and the intracellular localization of the polyclonal antibody was investigated using immunofluorescence assay (IFA). Results SDS-PAGE showed successful construction of the recombinant expression plasmid, and Coomassie brilliant blue staining showed the generation of the high-purity recombinant GRA24 truncated protein. ELISA measured that the titer of the polyclonal antibody against the GRA24 truncated protein was higher than 1:208 400, and Western blotting showed that the polyclonal antibody was effective to recognize the endogenous GRA24 proteins of different T. gondii strains and specifically recognize the recombinant GRA24 truncated protein. Indirect IFA showed that the GRA24 protein secreted 16 hour following T. gondii invasion in host cells. Conclusions The polyclonal antibody against the T. gondii GRA24 protein has been successfully prepared, which has a widespread applicability, high titers and a high specificity. This polyclonal antibody is available for Western blotting and IFA, which provides the basis for investigating the function of the GRA24 protein.
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Objective To establish a low density, high purity and high stability in vitro culture method of primary hippocampal neurons of fetal rats by co-culturing hippocampal and cortical cells, so as to obtain higher purity and better vitality of primary hippocampal neurons disease. Methods The fetal rat hippocampal tissue was isolated from 16-18 days pregnant SD rats, then cut and digested by 0.125% trypsin. The obtained cell suspension was filtered by 200 mesh cell sieve, and then the obtained cell suspensions were then inoculated into the inner layer and outer ring of the culture plate in a surrounding form. They were co-cultured in DMEM/F12 medium containing 10% horse serum. After 4-6 hours of cell adhesion, the culture medium was changed to maintenance medium (Neurobasal+2% B27+0.5 mmol/L glutamine). Then the cell viability was detected with CCK-8 kit and the purity of hippocampal neurons was detected by immunofluorescent staining. Results Hippocampal neurons grew well and formed crisscross neural networks after 5 days. And it could survive for 3 weeks. The purity of hippocampal neurons was up to 98%. Conclusion The method of co-culturing hippocampal and cortical cells can obtain high-purity, high activity, high survival rate, and high stability primary hippocampal neurons from fetal rats, which can provide certain experimental conditions for the study of hippocampal neuron related diseases in the nervous system and is worthy of promotion and application.
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Objective Optimizing the preparation method to improve the quality of mouse lung cryosections to help enhance the specificity of immunofluorescence staining and obtain more accurate and reliable experimental result.Methods C57BL/6 mouse lung tissue was used to make cryosections via the traditional post-freezing fixation method,pre-freezing fixation method,and a modified perfusion pre-freezing fixation method.A laser-scanning confocal microscope was used to observe lung tissue immunofluorescence staining.Whole areas of mouse lung slices were scanned by fluorescence microscope,and the numbers of intact airways per unit area of lung tissue were calculated.Results In the lung cryosections made via the traditional post-freezing fixation method,the alveoli structure was damaged,the airway wall was seriously disrupted,and there was non-specific staining.Lung cryosections made via the pre-freezing fixation method showed relatively intact alveolar and airway structures but collapsed alveoli and several destroyed airways.In the lung cryosections obtained via the modified perfusion pre-freezing fixation method,the structure and morphology of the alveoli and airways were intact and clear.Additionally,the locations of multiple proteins targeted with immunofluorescence staining were accurate.The number of intact airways(diameter ≥100 μm)per unit area in the lung cryosections obtained via the modified perfusion pre-freezing fixation method was higher than that from cryosections made using the pre-freezing fixation method((0.66±0.15)/mm2 vs(0.33±0.14)/mm2,P＜0.05)and was also significantly higher than that from sections made using the traditional post-freezing fixation method((0.66±0.15)/mm2 vs(0.02±0.04)/mm2,P＜0.01).Conclusions The modified perfusion pre-freezing fixation method for cryosections is conducive to maintaining the integrity of mouse lung tissue morphology and obtaining high-quality multiplex immunofluorescence staining result.
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Abstract High expression of MMP-2 and MMP-9 in periapical lesions plays an important role in the degradation of the extracellular matrix. This study aimed to investigate the effect of epigallocatechin-3-gallate (EGCG)-based endodontic paste as an intracanal dressing on the expression of MMP-2 and MMP-9 in periapical lesions. Periapical lesions were experimentally induced in 35 mature beagle dog premolars randomly divided into healthy teeth, untreated periapical lesions, periapical lesions treated in a single session (control groups), and periapical lesions treated in two sessions with EGCG or calcium hydroxide-based pastes (experimental groups). After 120 days, specimens were obtained for histopathologic and immunofluorescence analyses to assess the expression of MMP-2 and MMP-9. The statistical analysis was performed using a p-value of 0.05. Endodontic treatment in two sessions using medication with EGCG and calcium hydroxide-based pastes provided similar repair of the apical and periapical tissues and neoformation of periodontal ligament fibers, cementum, and alveolar bone (p>0.05). The experimental groups treated in two sessions with both medications presented expression of MMP-2 and MMP-9 similar to that in healthy teeth (p>0.05), and significantly lower than teeth treated in a single session or untreated periapical lesions (p <0.001). Expression of MMP-2 and MMP-9 was observed in the cytoplasm of fibroblasts, osteoblasts, cementoblasts, cementocytes, and vascular endothelium. The use of EGCG-based endodontic paste reduced the expression of MMP-2 and MMP-9 and allowed repair of periapical lesions, similar to calcium hydroxide-based paste, and superior to treatment performed in a single session.
Resumo A alta expressão de MMP-2 e MMP-9 em lesões periapicais desempenha um papel importante na degradação da matriz extracelular. Este estudo teve como objetivo investigar o efeito de uma pasta à base de epigalocatequina-3-galato (EGCG) como curativo intracanal na expressão de MMP-2 e MMP-9 em lesões periapicais. Lesões periapicais foram induzidas experimentalmente em 35 pré-molares de cães da raça beagle, maduros, divididos aleatoriamente em dentes saudáveis, lesões periapicais não tratadas, lesões periapicais tratadas em uma única sessão e lesões periapicais tratadas em duas sessões com a pasta de EGCG ou pasta à base de hidróxido de cálcio. O operador monitorou os animais e realizou a eutanásia após 120 dias para análises histopatológicas e de imunofluorescência para avaliar a expressão de MMP-2 e MMP-9. A análise estatística foi realizada utilizando p=0,05. O tratamento endodôntico em duas sessões com pasta à base de EGCG e pasta à base de hidróxido de cálcio proporcionou níveis semelhantes de reparação dos tecidos apicais e periapicais e neoformação de fibras do ligamento periodontal, cemento e osso alveolar. Em ambos os grupos, a expressão de MMP-2 e MMP-9 foi mínima, sendo observada no citoplasma de fibroblastos, osteoblastos, cementoblastos, cementócitos e endotélio vascular. Em ambos os grupos tratados em duas sessões, a expressão de MMP-2 e MMP-9 foi semelhante à dos dentes hígidos e significativamente menor do que nas lesões periapicais tratadas em sessão única ou não tratadas (p < 0,001). O uso da pasta à base de EGCG reduziu a expressão de MMP-2 e MMP-9 e permitiu o reparo de lesões periapicais, semelhante à pasta à base de hidróxido de cálcio, e foi superior ao tratamento realizado em sessão única.
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Objectives: To detect the diagnostic accuracy of rapid antibody detection test using IgM immunochromatography for scrub typhus in children. Methods: This cross-sectional study enrolled children aged 2 months-18 years hospitalized over a period of 18 months with undifferentiated fever of duration five days or more. The blood samples were subjected to serological tests like Weil-Felix, Scrub IgM ELISA, immunofluroscence assay (IFA) and rapid diagnostic test (IgM Immunochromatography). Diagnostic accuracy was measured against IFA as the gold standard. Results: A total of 90 children were included in the study, among which 43 children were positive for gold standard test IFA. Rapid diagnostic test showed sensitivity of 88.3%, specificity of 89.3%, positive predictive value of 88.3% and negative predictive value of 89.3%. The sensitivity, specificity, PPV and NPV of Weil-Felix test was 39.5%, 84.2%, 58.6 and 71.1%, respectively and of IgM ELISA was 93%, 89.3%,88.8% and 93.3%, respectively. Conclusion: IgM immunochromatography had good diagnostic accuracy for scrub typhus in children with acute undifferentiated fever.
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Background & objectives: The diagnosis of scrub typhus (ST) is usually done using enzyme-linked immunosorbent assay (ELISA) due to its ease of performance and reading objectivity. The cut-off value for ELISA needs to be calculated for each geographical location as it depends on zonal endemicity of the disease. This study was, therefore, undertaken to calculate the pan-India cut-off for anti-Orientia tsutsugamushi (OT) immunoglobulin M (IgM) by ELISA. Methods: Samples from cases (cases of ST) and controls (voluntary, consenting, healthy adults) were collected by a network of 29 laboratories across India and tested for anti-OT IgM by immunofluorescence assay (IFA), the considered gold standard test. These samples were retested by ELISA for anti-OT IgM and their optical densities (ODs) were used for cut-off estimation by receiver operating characteristic (ROC) curve. Results: Anti-OT IgM ELISA ODs from 273 controls and 136 cases were used for the cut-off estimation. The ODs of the anti-OT IgM ELISA on healthy individuals and those of confirmed ST cases ranged from 0.1 to 0.75 and 0.5 to 4.718, respectively. ROC curve-based cut-off for ELISA was calculated as 0.554 at a sensitivity of 95.2 per cent and specificity of 95.1 per cent. A value of >1 was noted to have a specificity of 100 per cent in diagnosing ST. Interpretation & conclusions: The cut-off calculated for India was similar to the previous cut-off that was used until now.
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Introduction: Diseases result from abnormal divergence of the normal structural and functional well-being of an organism. It can be brought about by physical, biological, chemical, genetic, or autoimmune causes. Autoimmune diseases occur when the body’s defence system targets its own healthy cells and tissues. The clinical signs and symptoms vary depending on the target tissues. Oral lesions such as ulcers, blisters, mucositis, and gingivitis are seen in many autoimmune diseases and may be an early sign, first recognized by the dental surgeon. Objective: To review the various autoimmune diseases affecting the orofacial region and update the clinicians about their oral manifestations. Materials and Methods: Case reports, review articles and original research papers published in various electronic databases like PubMed, Cross reference, Google scholar, and data collected from books are compiled in this review article. Result and Conclusion: This review gives an overview of some of the common autoimmune diseases affecting the head and neck region, their pathogenesis, clinical features, histopathological features and laboratory findings.
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Deacetylation of chitin is closely related to insect development and metamorphosis. Chitin deacetylase (CDA) is a key enzyme in the process. However, to date, the CDAs of Bombyx mori (BmCDAs), which is a model Lepidopteran insect, were not well studied. In order to better understand the role of BmCDAs in the metamorphosis and development of silkworm, the BmCDA2 which is highly expressed in epidermis was selected to study by bioinformatics methods, protein expression purification and immunofluorescence localization. The results showed that the two mRNA splicing forms of BmCDA2, namely BmCDA2a and BmCDA2b, were highly expressed in the larval and pupal epidermis, respectively. Both genes had chitin deacetylase catalytic domain, chitin binding domain and low density lipoprotein receptor domain. Western blot showed that the BmCDA2 protein was mainly expressed in the epidermis. Moreover, fluorescence immunolocalization showed that BmCDA2 protein gradually increased and accumulated with the formation of larval new epidermis, suggesting that BmCDA2 may be involved in the formation or assembly of larval new epidermis. The results increased our understandings to the biological functions of BmCDAs, and may facilitate the CDA study of other insects.
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Animals , Bombyx/metabolism , Metamorphosis, Biological/genetics , Larva/metabolism , Gene Expression , Insect Proteins/metabolism , Chitinالملخص
Somatostatin (SST) is an inhibitory polypeptide hormone that plays an important role in a variety of biological processes. Somatostatin receptor 2 (SSTR2) is the most widely expressed somatostatin receptor. However, the specific cell types expressing Sstr2 in the tissues have not been investigated. In this study, we detected the expression pattern of SSTR2 protein in mouse at different development stages, including the embryonic 15.5 days and the postnatal 1, 7, 15 days as well as 3 and 6 months, by multicolour immunofluorescence analyses. We found that Sstr2 was expressed in some specific cells types of several tissues, including the neuronal cells and astrocytes in the brain, the mesenchymal cells, the hematopoietic cells, the early hematopoietic stem cells, and the B cells in the bone marrow, the macrophages, the type Ⅱ alveolar epithelial cells, and the airway ciliated cells in the lung, the epithelial cells and the neuronal cells in the intestine, the hair follicle cells, the gastric epithelial cells, the hematopoietic stem cells and the nerve fibre in the spleen, and the tubular epithelial cells in the kidney. This study identified the specific cell types expressing Sstr2 in mouse at different developmental stages, providing new insights into the physiological function of SST and SSTR2 in several cell types.
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Mice , Animals , Receptors, Somatostatin/metabolism , Hematopoietic Stem Cells/metabolism , Epithelial Cellsالملخص
Introduction@#Epidermolysis Bullosa Acquisita (EBA) is a rare autoimmune blistering disease which presents in the skin and mucous membranes. The decrease in anchoring fibrils in the basement membrane zone causes separation of the epidermis from the dermis, resulting in its blistering presentation. The treatment plan will depend on the severity of the disease. The first-line treatment for mild EBA includes topical corticosteroids and immunomodulators such as dapsone and colchicine; while severe cases of EBA may be given intravenous immunoglobulins, systemic steroids, and immunosuppressants such as azathioprine and cyclophosphamide. @*Case Report@#This is a case of a 50-year-old Filipino male who presented with a 2-year history of vesicles and tense bullae which evolved into papules, plaques and erosions with scarring and milia formation on the scalp and trauma-prone areas of the trunk and extremities. Clinical examination revealed multiple, well-defined, irregularly shaped erythematous papules and plaques with crusts, scales, erosions, pearl-like milia and scarring on the chest, back, upper, and lower extremities. The oral mucosa was moist with some ulcers on the tongue. Histopathologic examination using Hematoxylin and Eosin (H&E) stain revealed the absence of the epidermis with retention of dermal papillae suggestive of subepidermal clefting. Further examination with direct immunofluorescence (DIF) revealed monoclonal immunoglobulin (IgG) deposits demonstrating an intense linear fluorescent band at the dermoepidermal junction, consistent with Epidermolysis Bullosa Acquisita. Overall, the combined administration of prednisone, azathioprine, and colchicine resulted only in transient and incomplete resolution of lesions in this case of EBA.@*Conclusion@#The management of EBA is mostly supportive with the goal of minimizing complications. Combination treatments using steroids, colchicine, and azathioprine have been reported with various results. Its management remains challenging as most cases are refractory to treatment.
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Epidermolysis Bullosa Acquisita , Azathioprine , Colchicine , Prednisoneالملخص
【Objective】 To observe the penetration and biological activity of PTD4-Cu, Zn-SOD into human astrocytes and whether it can mitigate hypoxia damages. 【Methods】 ①Immunohistochemistry and fluorescence test: We labeled the Cu, Zn-SOD by a monoclonal antibody, combined it with the fluorescent secondary antibody labeled with fluorescein isothiocyanate (FITC) to observe the effect of transduced PTD4-Cu, Zn-SOD on the viability of human astrocytes. ② The experimental group: After hypoxic damage model, the cells were divided into three groups: blank control, group Cu, Zn-SOD, and group PTD4-Cu, Zn-SOD. Group blank was added with DMEM medium (excluding serum) as control; DMEM medium was added to the other two for one hour (excluding serum) with its fusion proteins (Cu, Zn-SOD and PTD4 -Cu, Zn-SOD) with the final concentration of 2 μmoL/L. After the intervention, we used SOD and MDA test kits to observe PTD4-Cu, Zn-SOD and Cu, Zn-SOD in astrocytes after fusion protein intervention. 【Results】 The PTD4-Cu, Zn-SOD fusion protein could have aggregation distribution in the nucleus by FITC fluorescently labeled. After the intervention, it could increase the SOD activity in astrocytes in group PTD4-Cu, Zn-SOD and group Cu, Zn-SOD compared with control group, but the SOD activity was more obvious in the fusion proteins PTD4-Cu, Zn-SOD group. And the dose of MDA was reduced in group PTD4-Cu, Zn-SOD compared with group Cu, Zn-SOD and control group. 【Conclusion】 PTD4-Cu, Zn-SOD fusion protein can transcellular membrane of human astrocytes. The fusion protein PTD4-Cu, Zn-SOD can increase the SOD activity and reduce the content of MDA by human astrocytes from hypoxia injury.
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Objective To investigate the protective effect and mechanism of liraglutide on the paraquat (PQ)⁃ induced Parkinson's disease (PD) mouse model. Methods Totally 24 Kunming mice were randomly divided into control group, PQ group and PQ +liraglutide group, 8 mice in each group. PD model was established by intraperitoneal injection of PQ (10 mg/kg) for 5 consecutive days, and liraglutide (50 nmol/kg) was injected intraperitoneally for 7 consecutive days. The free⁃standing and locomotor activity of mice were measured by behavioral method. Immunofluorescence was used to observe the number of tyrosine hydroxylase (TH) and ionized calcium binding adaptor molecule 1 (Iba1) immunoreactive cells. Western blotting was used to detect the expression of protein TH, glial fibrillary acidic protein (GFAP), mitofusin⁃2 (Mfn2) and dynamin⁃related protein 1 (Drp1). Results The numbers of free⁃standing and locomotor activity numbers decreased significantly (P<0.01, P < 0.05) in PQ group compared with the control group, and the number of TH immunoreactive cells and TH protein expression in substantia nigra decreased significantly (P<0.01, P<0.01) compared with the control group, while the number of Iba1 immunoreactive cells and GFAP protein expression increased significantly (P<0.01, P<0.01) compared with the control group; the expression of Drp1 protein in PQ group was significantly higher than that in control group (P<0.05), while the Mfn2 protein expression decreased significantly (P<0.05) compared with the control group. After treatment with liraglutide, the number of TH positive cells in PQ + liraglutide group was significantly lower than that in control group (P<0.05); the numbers of free⁃standing and locomotor activity increased significantly (P<0.05, P<0.05) in PQ + liraglutide group compared with the PQ group, and the number of TH positive cells and expression of TH protein in PQ + liraglutide group were significantly higher than that in PQ group (P<0.01, P< 0.01); while the number of Iba1 positive cells and GFAP protein expression decreased significantly (P<0.01, P<0.05) compared with the PQ group; the Drp1 protein expression decreased significantly (P<0.01) compared with the PQ group, while the expression of Mfn2 protein in PQ + liraglutide group was significantly higher than that in PQ group (P<0.01). Conclusion Liraglutide has neuroprotective effect by reducing neuroinflammation in substantia nigra, regulating mitochondrial fusion and fission.
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Objective To explore the effect of interleukin (IL)-6 on nucleated erythrocytes in lipopolysaccharide (LPS)-induced preeclampsia rats. Methods ELISA and immunohistochemistry were used to detect the IL-6 in peripheral blood and placenta of preeclampsia and normal pregnancy; Flow cytometry and immunofluorescence were used to detect the maternal nucleated erythrocytes. Pregnant SD rats were randomly divided into 3 groups: the control, LPS and LPS +anti-IL-6 group; IL-6, the proportion of nucleated erythrocytes, JAK2/MEK and PI3K/Akt signal-related genes were detected. Results The IL-6 of preeclampsia was higher than that of normal patients. Compared with the Control group, IL-6, the proportion of nucleated erythrocytes and JAK2, P85, Akt, P65, IL-IB mRNA of LPS group increased, the fetal weight decreased; Compared with the LPS group, IL-6, the proportion of nucleated erythrocytes and JAK2, P85, Akt, P65 and IL-IB mRNA of the LPS + anti-IL-6 group decreased. Conclusion The up-regulation of IL-6 of preeclampsia patients is accompanied by increased nucleated erythrocytes in peripheral blood. Neutralizing IL-6 in vivo may down-regulate JAK2/ PI3K/Akt/NF-KB-signal-mediated IL-IB to protect preeclampsia rats.
الملخص
Objective To investigate the structural distribution features and mechanism of elastic fibers and collagen fibers in ventricular interstitium of aged rats. Methods Five young SD rats (24 weeks) and five old SD rats (104 weeks) were used,and their cardiac function was examined by echocardiography. Modified Weigert elastic fiber staining, immunohistochemistry, immunofluorescence and Western blotting techniques were used to detect the expression changes of type I and IH collagen fibers and their proteins, elastic fibers and their proteins, matrix metalloproteinase 2 (MMP-2), matrix metalloproteinase 9 (MMP-9) and tissue inhibitor of metalloproteinase 2 (TIMP-2), respectively. Results The type I and type IH collagen in the ventricular interstitium of aged rats was very sufficient and wrapped around the cardiomyocytes. Compared with the young rats, the content of collagen protein in the ventricular interstitium of the aged rats significantly increased (P<0. 05). Elastic fibers in the ventricular interstitium of the aged rats were and widely distributed. Compared with the young rats, the number of elastic fibers and the level of elastin in the ventricular interstitium of the aged rats significantly decreased (P<0. 05), and the expression levels of MMP-2 and MMP-9 in ventricular muscle of aged rats increased, and the)' were correlated with the level of elastin. The level of TIMP-2 in ventricular muscle of aged rats decreased with age. Conclusion The number of collagen fibers and elastic fibers in ventricular interstitium of aged rats is fluctuated with each other. With the increase of age, the contents of TIMP-2 and elastic fibers in the ventricular interstitium gradually decreased, and the ratio of collagen fibers to elastic fibers is out of balance.