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Objective:To determine the expression of T follicular helper (Tfh) cell-related molecules inducible costimulator (ICOS) and programmed cell death-1 receptor (PD-1) in skin lesions of patients with bullous pemphigoid (BP), and to explore the role of Tfh cells in the pathogenesis of BP.Methods:Twenty-one paraffin-embedded tissue specimens were collected from 21 patients with confirmed BP in Dalian Dermatosis Hospital from 2014 to 2017, including 7 females and 14 males with an average age of 72.57 years. Ten normal skin tissue specimens served as control group. Immunohistochemical SP method was used to determine the expression of ICOS and PD-1 in BP skin lesions and normal skin tissues.Results:In BP lesions, ICOS and PD-1 were mainly expressed in the basal layer, spinous layer, granular layer and stratum corneum, especially in the spinous layer in the epidermis, and also expressed in inflammatory cells in the dermis. Additionally, they were both expressed in the cytoplasm and nuclei, occasionally expressed on the cell membrane. However, ICOS and PD-1 were rarely expressed in the normal skin tissues. The expression rates of ICOS and PD-1 were significantly higher in the BP group (85.71% [18/21], 47.62% [10/21], respectively) than in the normal control group (both were 0; P < 0.001, < 0.05, respectively) .Conclusion:Tfh cell-related molecules ICOS and PD-1 may play important roles in the pathogenesis of BP.
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Objective:To investigate the expression of integrin-linked kinase (ILK)/zinc finger transcription factor (Snail) signaling pathway in renal tubular epithelial-mesenchymal transition (EMT) in diabetic nephropathy (DN) and the effect of rhein.Methods:Healthy male Wistar rats of 8 weeks old were randomly divided into normal group, diabetic nephropathy group, rhein intervention group and valsartan intervention group, with 12 rats in each group. Streptozotocin (STZ) was used to induce the diabetic nephropathy model, then rhein intervention group and valsartan intervention group were given rhein 100 mg/(kg·d) and valsartan 30 mg/(kg·d), respectively. At the end of the 8th and 16th week, six rats of each group were killed, in situ lavage kidney, take out the kidney tissue of rats after fixed in wax block and slices. Renal tubular interstitial damage index and the relative area of interstitial collagen evaluated by hematoxylin-eosin (HE) and Masson staining respectively. The protein expression of E-cadherin, α-smooth muscle actin (α-SMA), ILK, Snail and matrix metalloproteinase-9 (MMP-2) in renal tubular epithelial cells were detected by immunohistochemistry.Results:Comparing to normal group, the renal tubular interstitial damage index and relative area of renal interstitial collagen of diabetic nephropathy rats were both increased. The expression of E-cadherin in renal tubular epithelial cells decreased and the expression of α-SMA significantly increased ( P<0.05). Comparing with diabetic nephropathy group, in rhein and valsartan intervention groups, the expression of E-cadherin in renal tubular epithelial cells increased, while the expression of α-SMA significantly decreased ( P<0.05). There was no significant difference in the above indexes between rhein intervention group and valsartan intervention group ( P>0.05). Compared to normal group, the expressions of ILK, Snail, MMP-2 increased progressively with the disease ( P<0.05). Compared with diabetic nephropathy group, rhein and valsartan intervention groups showed significant decrease in expression of ILK, Snail, MMP-2 ( P<0.05). There was no significant difference in the above indexes between rhein intervention group and valsartan intervention group ( P>0.05). Conclusions:Rhein could inhibit EMT progression by down-regulating the expression of ILK/Snail signaling pathway in renal tubular epithelial cells of DN rats.
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Objective To investigate the possible regulating effect of integrin-linked kinase (ILK) towards matrix metalloproteinase-9/tissue inhibitor of metalloproteinase-1 (MMP-9/TIMP-1) ratio in the process of transforming growth factor-β1 (TGF-β1)-induced epithelial-mesenchymal transition (EMT) in human kidney proximal tubular epithelial (HK-2) cells.Methods HK-2 cells were cultured and stimulated with 10 ng/ml TGF-β1.Specific ILK-small interfering RNA (ILK-siRNA) was used to inhibit ILK expression.The characteristic epithelial marker (E-cadherin) and mesenchymal marker (α-SMA) of EMT were examined by Real-time reverse transcription polymerase chain reaction (real-time RT-PCR) and Western blot.The expressions of ILK,MMP-9,and TIMP-1 were also examined,to determine the regulating effect of ILK towards MMP-9/TIMP-1 ratio.Results In the HK-2 cells cultured with TGF-β1,the expression of E-cadherin decreased,and α-SMA expression increased;overexpression of ILK and an abnormal changing of MMP-9/TIMP-1 ratio were observed.ILK inhibition by ILK-siRNA could adjust MMP-9/TIMP-1 ratio to near normal.Meanwhile,the overexpressed ILK and α-SMA were decreased.Conclusions Our data indicates that ILK-siRNA successfully inhibits ILK expression,which regulates the MMP-9/TIMP-1 ratio in HK-2 cells.The inhibition of ILK expression suppresses TGF-β1-induced EMT partially.
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Three cases of swimming pool granuloma are reported.Case 1:a 40-year-old female presented with a 2-month history of nodules and plaques on the right hand and forearm.She was a tropical fish salesperson but denied trauma history.Skin examination revealed multiple irregularly sized,dark-red nodules and plaques on the joints of right fingers,wrist,and elbow,as well as multiple subcutaneous nodules simulating strings of beads on the right upper limb.Case 2:a 48-year-old female presented with a 2-month history of nodules and plaques on the left hand and forearm.There was a history of trauma due to tropical fish tank and filter cleaning.Physical examination showed multiple deep purple plaques and painless subcutaneous nodules scattered on the left hand,wrist,and upper limb.Case 3:a 39-year-old male presented with a 3-month history of nodules on the fingers of both hands.There was no history of trauma,but he was a tropical aquarist.Skin examination revealed multiple soybean-sized dark-red nodules on the extensor aspect of interphalangeal joints of both hands.Fungal examinations yielded negative results in the 3 cases,while histopathology revealed infectious granuloma with a mixed inflammatory cell infiltrate.All of the cases showed positive results in purified protein derivative (PPD)skin test.Mycobacterium marinum was isolated from the lesional tissue of Case 1 and 2,but not from Case 3.All the patients were diagnosed with swimming pool granuloma,and given anti-atypical mycobacterial therapy including oral rifampin and clarithromycin.The lesions disappeared after 1 to 3 months of treatment,with the treatment course varying from 2 to 5 months.No recurrence was observed during a 3- to 12-month follow-up.
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Objective To establish an in vitro model for malignant melanoma with a malignant melanoma cell line MV3 on de-epidermized dermis (DED) and to study the invasion mode of melanoma cells. Methods A human de-epidermized dermis was prepared with some elements of basal membrane (BM). Then, the reconstructed BM was identified by periodic acid schiff (PAS) staining and immunochemical staining for collagen Ⅳ. MV3 cells were seeded onto the prepared acellular dermis and maintained at the air-liquid interface for 13-15 days after 3-day submerged culture. Subsequently, the reconstructed malignant melanoma tissue was examined with hematoxylin and eosin (HE) staining and immunohistochemical staining with antibodies to S-100 protein and HMB45. Results No obvious changes were observed by naked eye in DED after the inoculation with MV3 cells. PAS staining and immunochemical staining for collagen Ⅳ confirmed the presence of BM component on the surface of DED and in the cavity of skin appendages in DED. Histological examination and immunochemical staining revealed that on the BM zone, MV3 cells grew into irregularly sized clusters; in the .cavity of skin appendages, they attached onto the BM and aggregated into circular or bandlike shape; and at the lateral side of DED, they invasively and diffusely grew, broke through the BM and intruded into the surrounding tissues of DED. The reconstructed tissue was positive for S-100 protein and weakly positive for HMB45. Conclusions The in vitro model of malignant melanoma could be reconstructed by skin organ culture system. And, the experiment suggests that BM could affect the invasive growth pattern of malignant melanoma cells.
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Objective To construct tissue-engineered skin via in vitro inoculation of epidermal stem cells(ESCS)onto de-epidermized dermis.Methods Skin tissue was obtained from the foreskin of a healthy 6-year-old child.and keratinocytes were isolated by two-step trypsinization method followed by the collection of ESCS via rapid adhesion by collagen Ⅳ.The ESCS were identified by morphological observation and immunohistochemical staining with K19 and integrin β1.To construct tissue-engineered skin,selected ESCS were seeded onto the surface of de-epidermized dermis followed by a one-week culture immersed in the medium and a subsequent 4-week culture at the air-medium interface.The tissue-engqneered skin was evaluated with haematoxylin & eosin(HE)staining as well as keratin immunohistochemistry.Results Micro scopically,cultured ESCs showed a paving stone-like appearance and grew into colonies.Immunohistochemistry revealed that the ESCs were positive for integrin-β1 and keratin 19.After 5 weeks of culture,3-6 layers of epidermal cell were observed on the dermis with the formation of stratum corneum.Keratin protein was observed in the artificial epidermal skin.Conclusion Tissue-engineered skin is successfully constructed with epidermal stem cells and de-epidermized dermis in vitro.