Background: Aged skin exhibits visual alterations such as wrinkles, rough texture, pore dilation, and dull skin tone, as well as physiological aging, namely, decreased hydration and increased transepidermal water loss...Background: Aged skin exhibits visual alterations such as wrinkles, rough texture, pore dilation, and dull skin tone, as well as physiological aging, namely, decreased hydration and increased transepidermal water loss (TEWL). Recent advances in coherence tomography have also revealed that skin aging affects in vivo epidermal keratinocyte architecture. However, the interconnectivity between spatial architectural aging and visual/physiological aging parameters remains largely unknown. Purpose: To elucidate whether the tomographic keratinocyte architectural aging is correlated with visual and physiological skin aging parameters and to quantitatively evaluate the improvements of the architectural, visual, and physiological aging parameters by the daily treatment of the skin care formula containing Galactomyces Ferment Filtrate (GFF, 8X Pitera<sup>TM</sup>). Method: We measured the in vivo keratinocyte cellular architecture with two-photon stereoscopic tomography obtaining by-layer epidermal section images in 78 Asian females of various ages. Visual aging parameters were analyzed using a portable image capture system. Hydration and TEWL were also assessed. The anti-aging effects of GFF-containing skin moisturizer (SK-II LXP Cream<sup>TM</sup>) were also examined in two studies after twice-daily application for 2 (N = 35) and 4 (N = 32) weeks. Results: As for the keratinocyte cellular architecture, skin aging was significantly associated with decreased cell density and increased cell uniformity. These architectural aging parameters were significantly correlated with visual and physiological aging parameters, namely, rough texture, wrinkles, pore dilation, dull skin tone, dehydration, and increased TEWL. The strong interconnectivity allowed us to develop formulae to estimate the keratinocyte architecture from visual aging parameters. Moreover, twice-daily application of SK-II significantly improved the keratinocyte architecture associated with multiple skin aging visual and physiological parameters. Conclusion: Skin aging is a process involving mutual interconnections among epidermal keratinocyte cellular architecture, visual, and physiological parameters. The GFF-containing moisturizer SK-II effectively improves spatial architecture of keratinocytes in epidermis and these evaluated skin aging parameters in a new trajectory over the course of treatment. .展开更多
BACKGROUND Current evidence shows that human induced pluripotent stem cells(hiPSCs)can effectively differentiate into keratinocytes(KCs),but its effect on skin burn healing has not been reported.AIM To observe the eff...BACKGROUND Current evidence shows that human induced pluripotent stem cells(hiPSCs)can effectively differentiate into keratinocytes(KCs),but its effect on skin burn healing has not been reported.AIM To observe the effects of hiPSCs-derived KCs transplantation on skin burn healing in mice and to preliminarily reveal the underlying mechanisms.METHODS An analysis of differentially expressed genes in burn wounds based on GEO datasets GSE140926,and GSE27186 was established.A differentiation medium containing retinoic acid and bone morphogenetic protein 4 was applied to induce hiPSCs to differentiate into KCs.The expression of KCs marker proteins was detected using immunofluorescence staining.A model of a C57BL/6 mouse with deep cutaneous second-degree burn was created,and then phosphate buffered saline(PBS),hiPSCs-KCs,or hiPSCs-KCs with knockdown of COL7A1 were injected around the wound surface.The wound healing,re-epithelialization,engraftment of hiPSCs-KCs into wounds,proinflammatory factor level,and the NF-κB pathway proteins were assessed by hematoxylin-eosin staining,carboxifluorescein diacetate succinimidyl ester(CFSE)fluorescence staining,enzyme linked immunosorbent assay,and Western blotting on days 3,7,and 14 after the injection,respectively.Moreover,the effects of COL7A1 knockdown on the proliferation and migration of hiPSCs-KCs were confirmed by immunohistochemistry,EdU,Transwell,and damage repair assays.RESULTS HiPSCs-KCs could express the hallmark proteins of KCs.COL7A1 was down-regulated in burn wound tissues and highly expressed in hiPSCs-KCs.Transplantation of hiPSCs-KCs into mice with burn wounds resulted in a significant decrease in wound area,an increase in wound re-epithelialization,a decrease in proinflammatory factors content,and an inhibition of NF-κB pathway activation compared to the PBS group.The in vitro assay showed that COL7A1 knockdown could rescue the inhibition of hiPSCs-KCs proliferation and migration,providing further evidence that COL7A1 speeds up burn wound healing by limiting cell proliferation and migration.CONCLUSION In deep,second-degree burn wounds,COL7A1 can promote KC proliferation and migration while also suppressing the inflammatory response.展开更多
Objective Psoriasis is often combined with metabolic abnormalities,such as obesity and diabetes.The upregulation of chemerin,which is an essential protein produced primarily from white fat,is strongly correlated to th...Objective Psoriasis is often combined with metabolic abnormalities,such as obesity and diabetes.The upregulation of chemerin,which is an essential protein produced primarily from white fat,is strongly correlated to the development of psoriasis.However,there is no clarification on its exact function and mechanism in disease pathogenesis.The present study aims to determine its function and mechanism in disease pathogenesis.Methods The present study used a psoriasislike inflammatory cell model and imiquimod(IMQ)-induced mouse model to confirm whether chemerin is upregulated in psoriasis patients.Results Chemerin enhanced the keratinocyte proliferation,inflammatory cytokine secretion,and activation of the MAPK signaling pathway.Crucially,the intraperitoneal injection of neutralizing anti-chemerin antibody(ChAb)diminished the epidermal proliferation and inflammation in the IMQ-induced mouse model.Conclusion The present results indicate that chemerin promotes keratinocyte proliferation,and enhances the production of inflammatory cytokines,thereby aggravating the psoriasis.Thus,chemerin can be a prospective target for the treatment of psoriasis.展开更多
Background: Sugar moiety of macromolecules is today very well known for its implications in many biological recognition mechanisms including cell-cell, extracellular matrix-cell and/or bacteria-cell interactions. In t...Background: Sugar moiety of macromolecules is today very well known for its implications in many biological recognition mechanisms including cell-cell, extracellular matrix-cell and/or bacteria-cell interactions. In this context lectins, which are carbohydrate-binding proteins displaying a high affinity for sugar groups of other molecules, are of a great importance, notably in immune response involving bacteria, viruses and fungi. As protein-carbohydrate interactions are often mediated by ions such as calcium, zinc or magnesium, we were prompted to study the effect of a thermal spring water (which contains this type of component) on interactions existing between: 1) osidic receptors of human normal keratinocytes and 2) two lectins greatly implicated in the immune response mechanisms (i.e. the dectin-1 and the langerin), and their ligands. Materials and Methods: In a first series of experiments, we studied the effect of increasing concentrations of a thermal spring water on interactions existing between glycosylated molecules and the osidic receptors expressed at the normal human keratinocytes surface. In a second step, and in order to better understand the putative effect of our thermal spring water on the immune response, we analyzed its effect on the interactions existing between the dectin-1 (implicated in the recognition of bacteria, viruses and fungi) and the langerin (expressed by Langerhans cells, the immune cells of the cutaneous tissue), and their ligands in a model using recombinant human lectins and appropriate binding molecules. Results: We showed here that our thermal spring water was able to reinforce interactions between keratinocytes osidic receptors and some of their ligands, in a dose-related manner: From 8% to 55% of increase with 10% to 30% (v/v) of thermal spring water. In the second part of our studies, we also showed that our thermal spring water was able to modulate interactions between dectin-1 and langerin and their ligands through a biphasic effect: Interactions were enhanced by more than 40% and 20% respectively with 10% of thermal spring water, and return to their basal level or lower for higher concentrations. Conclusion: The tested thermal spring water, probably due to its ionic composition, could significantly affect interactions of osidic receptors with their ligands. This property could be of a great interest to help immune system to maintain an appropriate “vigilance state” by using the thermal water at up to a concentration of 10%, and by avoiding any runaway reaction in case of aggression, by using concentrations higher than 10%. .展开更多
Using inducers to induce cells to produce inflammatory response is a common in vitro experimental method to study inflammation.However,there are relatively few inflammatory models developed for the cosmetic industry,a...Using inducers to induce cells to produce inflammatory response is a common in vitro experimental method to study inflammation.However,there are relatively few inflammatory models developed for the cosmetic industry,and there are also great differences in the control of model induction,the selection of inflammatory indicators,and the concentration of inducers.Therefore,in this paper,by systematically studying the effects of Lipopolysaccharide(LPS)on the cell viability,the levels of IL-1α,IL-8 and ROS of human primary keratinocytes,a skin inflammation model based human primary keratinocyte was developed.The results showed that 0.01~100μg/mL LPS had no significant effect on the cell viability of human primary keratinocytes,while 100μg/mL LPS could simultaneously induce human primary keratinocytes to produce large amounts of IL-1αand IL-8,and 0.01μg/mL LPS could induce plentiful ROS.Therefore,a skin inflammation model for differential induction of different inflammatory indicators was established,and the sample OSM2021041301 was tested with this model,it was found that sample OSM2021041301 could significantly inhibit LPS-induced elevated IL-1αand IL-8 levels,the inhibitory effect on LPS-induced elevated ROS level was weak.The results indicated that OSM2021041301 has certain anti-inflammatory effect on inflammation caused by the increase of IL-1α,IL-8 and ROS induced by LPS and its analogues.展开更多
Objective Psoriasis is an immune-mediated inflammatory disease.Despite advances in the study of its pathogenesis,the exact development mechanism of psoriasis remains to be fully elucidated.Hyperproliferative epidermis...Objective Psoriasis is an immune-mediated inflammatory disease.Despite advances in the study of its pathogenesis,the exact development mechanism of psoriasis remains to be fully elucidated.Hyperproliferative epidermis plays a crucial role in psoriasis.This study aimed to investigate the effects of interleukin-36β(IL-36β)on keratinocyte dysfunction in vitro.Methods Human keratinocyte cell lines,HaCaT cells,were treated with 0(control),50 or 100 ng/ml IL-36βrespectively for 24 h.Cell viability was determined with a cell counting kit-8 assay.Flow cytometry was used to assess the effects of IL-36βon apoptosis and cell cycle distribution.Expressions of the differentiation markers,such as keratin 10 and involucrin,were evaluated by quantitative real-time polymerase chain reaction(RT-qPCR).Expressions of the inflammatory cytokines,IL-1βand IL-6 were tested by ELISA.Results CCK8 assay showed the survival rate had no significant difference between the control and treated group(P>0.05).Flow cytometry analysis showed cell cycle arrest at S phase in the IL-36β-treated groups compared with the control group(P<0.05).RT-qPCR verified the decreased mRNA expressions of keratin 10 and involucrin in the IL-36β-treated groups compared with the negative control(P<0.01).ELISA showed 100 ng/ml IL-36βenhanced levels of IL-1βand IL-6 in culture supernatants of HaCaT cells compared with the negative control(P<0.05).Conclusion Taken together,these findings suggest that IL-36βcould induce cell cycle arrest at S phase,inhibit keratin 10 and involucrin expressions and promote inflammatory activity in HaCaT cell lines.展开更多
AIM:To investigate the healing process after severe corneal epithelial damage in rats treated with mesenchymal stem cells(MSCs)cultured with or without keratinocyte growth factor(KGF-2)and autologous serum(AS)on amnio...AIM:To investigate the healing process after severe corneal epithelial damage in rats treated with mesenchymal stem cells(MSCs)cultured with or without keratinocyte growth factor(KGF-2)and autologous serum(AS)on amniotic membrane(AM).Many patients are blind and devastated by severe ocular surface diseases due to limbal stem cell deficiency.Bone marrow-derived MSCs are potential sources for cellbased tissue engineering to repair or replace the corneal tissue,having the potential to differentiate to epithelial cells.METHODS:The study included 5 groups each including 10 female'Sprague Dawley'rats in addition to20 male rats used as bone marrow donors.Group I rats received AM+MSCs,Group II rats AM+MSCs cultured with KGF-2,Group III rats AM+MSCs cultured with KGF-2+AS,Group IV rats only AM and Group V rats,none.AS was derived from blood drawn from male rats and bone marrow was obtained from the femur and tibia bones of the same animals.Therapeutic effect was evaluated with clinical,histopathological and immunohistochemical assessment.MSC engraftment was demonstrated via detection of donor genotype(Y+)in the recipient tissue(X)with polymerase chain reaction.RESULTS:Corneal healing was significantly better in Groups I-III rats treated with MSC transplantation compared to Group IV and Group V rats with supportive treatment only.The best results were obtained in Group III rats with 90%transparency,70%lack of neovascularization,and 100%epithelium damage limited to less than 1/4 of cornea.CONCLUSION:We suggest that culture of MSCs with KGF-2 and AS on AM is effective in corneal repair in case of irreversible damage to limbal stem cells.展开更多
AIM:To investigate the effect of keratinocyte growth factor(KGF) gene therapy in acetic acid-induced ulcerative colitis in rat model.METHODS:The colitis of Sprague-Dawley rats was induced by intrarectal infusion of 1 ...AIM:To investigate the effect of keratinocyte growth factor(KGF) gene therapy in acetic acid-induced ulcerative colitis in rat model.METHODS:The colitis of Sprague-Dawley rats was induced by intrarectal infusion of 1 mL 5%(v/v) acetic acid.Twenty-four hours after exposed to acetic acid,rats were divided into three experimental groups:control group,attenuated Salmonella typhimurium Ty21a strain(SP) group and SP strain carrying human KGF gene(SPK) group,and they were separately administered orally with 10% NaHCO3,SP or SPK.Animals were sacrificed and colonic tissues were harvested respectively on day 3,5,7 and 10 after administration.Weights of rats,colonic weight/length ratio and stool score were evaluated.Histological changes of colonic tissues were examined by hematoxylin and eosin(HE) staining method.The expression of KGF,KGF receptor(KGFR) and TNF-α were measured either by enzyme-linked immunosorbent assay or Western blotting.Immunohistochemistry was used to detect the cellular localization of KGFR and Ki67.In addition,superoxide dismutase(SOD) activity and malondialdehyde(MDA) contents in the homogenate were measured.RESULTS:Body weight and colonic weight/length ratio were declined in SPK group compared with SP and control groups(body weight:272.78 ± 17.92 g vs 243.72 ± 14.02 g and 240.68 ± 12.63 g,P < 0.01;colonic weight/length ratio:115.76 ± 7.47 vs 150.32 ± 5.99 and 153.67 ± 5.50 mg/cm,P < 0.01).Moreover,pathological changes of damaged colon were improved in SPK group as well.After administration of SPK strain,KGF expression increased markedly from the 3rd d,and remained at a high level till the 10th d.Furthermore,KGFR expression and Ki67 expression elevated,whereas TNF-α expression was inhibited in SPK group.In the group administered with SPK,SOD activity increased significantly(d 5:26.18 ± 5.84 vs 18.12 ± 3.30 and 18.79 ± 4.74 U/mg,P < 0.01;d 7:35.48 ± 3.35 vs 22.57 ± 3.44 and 21.69 ± 3.94 U/mg,P < 0.01;d 10:46.10 ± 6.23 vs 25.35 ± 4.76 and 27.82 ± 6.42 U/mg,P < 0.01) and MDA contents decreased accordingly(d 7:7.40 ± 0.88 vs 9.81 ± 1.21 and 10.45 ± 1.40 nmol/mg,P < 0.01;d 10:4.36 ± 0.62 vs 8.41 ± 0.92 and 8.71 ± 1.27 nmol/mg,P < 0.01),compared with SP and control groups.CONCLUSION:KGF gene therapy mediated by attenuated Salmonella ameliorates ulcerative colitis induced by acetic acids,and it may be a safe and effective treatment for ulcerative colitis.展开更多
Objective:To examine the level of expression of anti skin aging gene of Ganoderma lucidum polysacchayides and clarify its mechanism with anti aging of this ancient Chinese medicine.Methods:HacaT cell of keratinocytes ...Objective:To examine the level of expression of anti skin aging gene of Ganoderma lucidum polysacchayides and clarify its mechanism with anti aging of this ancient Chinese medicine.Methods:HacaT cell of keratinocytes lines were cultured and treated with the polysaccharides.The total RNA was extracted with Trizol reagent and cDNA was synthesized by reverse thanscription.The obtained cDNAs were then fluorescently labeled with cy3 and cy5 respectively and hybridized with gene expressing pedigree cDNA chip.The images were scanned and analyzed with special software.The scan data were analyzed with software and checked by real time PCR.Results:Among total 18 346 human genes,the expression of 103 ones was up-regulated and 378 ones down-regulated.It was demonstrated evidently that Ganoderma lucidum polysaccharides affected the expression of genes of anti skin aging.Two ways are anastomotic.Conclusion:it is concluded by analysis of function of these up-regulation and down-regulation genes that Ganoderma lucidum polysaccharides may play an important role in boosting cell growth and against skin aging.It shows that the results of gene array reliable by real time PCR.展开更多
Objective:To investigate the effect of lead on the production of IL-8 and cell proliferation in normal human oral keratinocytes(NHKs).Methods:NHKs were prepared as outgrowths from normal human buccal mucosa.The cell...Objective:To investigate the effect of lead on the production of IL-8 and cell proliferation in normal human oral keratinocytes(NHKs).Methods:NHKs were prepared as outgrowths from normal human buccal mucosa.The cells were treated with three concentrations of lead glutamate(4.5×10<sup>-5</sup>M,4.5×10<sup>-6</sup>M and 4.5×10<sup>-7</sup>M).NHKs grown in glutamic acid were used as control.The amounts of IL-8 secreted in the culture supernatants were evaluated at 12 and 24 h using enzyme-linked immunospecific assay(ELISA).Cell proliferation was determined by the MTT colorimetric assay.Three cultures were used for each experiment,and three independent experiments were performed.Analysis of variance and Duncan’s multiple range tests were used for statistical analysis.Results:An elevation of IL-8 in culture supernatants of NHKs treated with lead at all concentrations at 12 and 24 h after exposure in a dose-dependent manner was revealed.A significant increase in cell numbers was observed only at 24 h exposed to 4.5×10<sup>- 5</sup>M lead glutamate.Conclusions:The capacity of NHKs,to secrete IL-8,enhanced by lead glutamate,is demonstrated here.Induction of cell proliferation is revealed only after exposure to high lead concentration.The elevation of secreted IL-8 is a probable initial sign for the acute inflammatory response and may be involved in the pathogenesis of lead stomatitis.展开更多
Liopxin A4(LXA4) is considered to be a crucial modulator in the inflammatory responses. In the present study, we aimed to study the effect of LXA4 on the inflammatory cytokines production induced by lipopolysacchar...Liopxin A4(LXA4) is considered to be a crucial modulator in the inflammatory responses. In the present study, we aimed to study the effect of LXA4 on the inflammatory cytokines production induced by lipopolysaccharide(LPS) and the possible mechanism in normal human epidermal keratinocytes(NHEKs). NHEKs were isolated and cultured. The expression of toll-like receptor 4(TLR4), LXA4 receptor(ALXR) and aryl hydrocarbon receptor(Ah R) in NHEKs was detected by reverse transcription polymerase chain reaction(RT-PCR). The m RNA and protein levels of tumor necrosis factor-alpha(TNF-α) and interleukin-1β(IL-1β) were determined in NHEKs stimulated by LPS(10 μg/m L) with or without preincubation with LXA4(100 nmol/L) for 30 min by real-time quantitative PCR(real-time q PCR) and enzyme-linked immunosorbent assay(ELISA), respectively. The expression levels of tumor necrosis factor receptor-associated factor 6(TRAF6) and suppressors of cytokine signaling 2(SOCS2) m RNAs and proteins, and nuclear translocation of NF-k B-p65 were measured by real-time q PCR and Western blotting, respectively. The results showed that NHEKs expressed TLR4, ALXR and Ah R. LXA4 significantly inhibited the m RNA and protein expression levels of TNF-α, IL-1β and TRAF6 induced by LPS in NHEKs, and LXA4 obviously increased the expression of SOCS2 at m RNA and protein levels. The nuclear NF-k B-p65 protein expression induced by LPS was inhibited after preincubation with LXA4 in NHEKs. It was concluded that LXA4 inhibits the LPS-induced production of TNF-α and IL-1β in NHEKs by up-regulating SOCS2 and down-regulating TRAF6.展开更多
The most effective sequence of small interfering RNA(si RNA) silencing STAT3 of psoriatic keratinocytes(KCs) was screened out,and the effects of the most effective si RNA combined with ultrasonic irradiation and S...The most effective sequence of small interfering RNA(si RNA) silencing STAT3 of psoriatic keratinocytes(KCs) was screened out,and the effects of the most effective si RNA combined with ultrasonic irradiation and Sono Vue microbubbles on the expression of STAT3 of KCs and the dose-and time-response were investigated.Three chemically-synthetic si RNAs targeting STAT3 carried by Lipofectamine 3000 were transfected into KCs,and the effects on STAT3 expression were detected,then the most effective si RNA was selected for the subsequent experiments.The negative controls of siR NA(si RNA-NC) labeled with Cy3 carried by Lipofectamine 3000 combined with ultrasonic irradiation and Sono Vue microbubbles were transfected into KCs,then the optimal parameters of ultrasonic irradiation were determined.The most effective si RNA carried by Lipofectamine 3000 combined with ultrasonic irradiation at the optimal parameters and Sono Vue microbubbles was transfected into KCs,and the dose-and time-response of RNA interference was determined.The effect of RNA interference by the most effective si RNA at the optimal time and dose carried by Lipofectamine 3000 combined with ultrasonic irradiation and Sono Vue microbubbles(LUS group) was compared with that only carried by Lipofectamine 3000(L group).The results showed that si RNA-3 achieved the highest silencing efficacy.0.5 W/cm2 and 30 s were selected as the parameters of ultrasonic irradiation.The si RNA-3 carried by Lipofectamine 3000 combined with ultrasonic irradiation and Sono Vue microbubbles could effectively knock down the STAT3 expression at m RNA and protein levels in dose-and time-dependent manners determined at 100 nmol/L with maximum downregulation on m RNA at 48 h,and on protein at 72 h after transfection.The LUS group achieved the highest silencing efficacy.It was concluded that si RNA-3 carried by Lipofectamine 3000 combined with ultrasonic irradiation and SonoV ue microbubbles could effectively knock down the STAT3 expression in psoriatic KCs,and the optimized transfection condition and the sequence of si RNA-3 could serve for further research on gene therapy of psoriasis.展开更多
文摘Background: Aged skin exhibits visual alterations such as wrinkles, rough texture, pore dilation, and dull skin tone, as well as physiological aging, namely, decreased hydration and increased transepidermal water loss (TEWL). Recent advances in coherence tomography have also revealed that skin aging affects in vivo epidermal keratinocyte architecture. However, the interconnectivity between spatial architectural aging and visual/physiological aging parameters remains largely unknown. Purpose: To elucidate whether the tomographic keratinocyte architectural aging is correlated with visual and physiological skin aging parameters and to quantitatively evaluate the improvements of the architectural, visual, and physiological aging parameters by the daily treatment of the skin care formula containing Galactomyces Ferment Filtrate (GFF, 8X Pitera<sup>TM</sup>). Method: We measured the in vivo keratinocyte cellular architecture with two-photon stereoscopic tomography obtaining by-layer epidermal section images in 78 Asian females of various ages. Visual aging parameters were analyzed using a portable image capture system. Hydration and TEWL were also assessed. The anti-aging effects of GFF-containing skin moisturizer (SK-II LXP Cream<sup>TM</sup>) were also examined in two studies after twice-daily application for 2 (N = 35) and 4 (N = 32) weeks. Results: As for the keratinocyte cellular architecture, skin aging was significantly associated with decreased cell density and increased cell uniformity. These architectural aging parameters were significantly correlated with visual and physiological aging parameters, namely, rough texture, wrinkles, pore dilation, dull skin tone, dehydration, and increased TEWL. The strong interconnectivity allowed us to develop formulae to estimate the keratinocyte architecture from visual aging parameters. Moreover, twice-daily application of SK-II significantly improved the keratinocyte architecture associated with multiple skin aging visual and physiological parameters. Conclusion: Skin aging is a process involving mutual interconnections among epidermal keratinocyte cellular architecture, visual, and physiological parameters. The GFF-containing moisturizer SK-II effectively improves spatial architecture of keratinocytes in epidermis and these evaluated skin aging parameters in a new trajectory over the course of treatment. .
基金Supported by the Hospital Research Fund,No.SDFEYBS1805,No.SDFEYGJ2013 and No.XKTJ-HRC20210015Suzhou Science and Technology Development Project,No.SYS2020105,No.SKJY2021078 and No.2022SS43+2 种基金the Special Project of“Technological Innovation”Project of CNNC Medical Industry Co.Ltd,No.ZHYLZD2021002CNNC Elite Talent Program2022 State Key Laboratory of Radiological Medicine and Radiation Protection jointly built by Province and Ministry,No.GZK1202244.
文摘BACKGROUND Current evidence shows that human induced pluripotent stem cells(hiPSCs)can effectively differentiate into keratinocytes(KCs),but its effect on skin burn healing has not been reported.AIM To observe the effects of hiPSCs-derived KCs transplantation on skin burn healing in mice and to preliminarily reveal the underlying mechanisms.METHODS An analysis of differentially expressed genes in burn wounds based on GEO datasets GSE140926,and GSE27186 was established.A differentiation medium containing retinoic acid and bone morphogenetic protein 4 was applied to induce hiPSCs to differentiate into KCs.The expression of KCs marker proteins was detected using immunofluorescence staining.A model of a C57BL/6 mouse with deep cutaneous second-degree burn was created,and then phosphate buffered saline(PBS),hiPSCs-KCs,or hiPSCs-KCs with knockdown of COL7A1 were injected around the wound surface.The wound healing,re-epithelialization,engraftment of hiPSCs-KCs into wounds,proinflammatory factor level,and the NF-κB pathway proteins were assessed by hematoxylin-eosin staining,carboxifluorescein diacetate succinimidyl ester(CFSE)fluorescence staining,enzyme linked immunosorbent assay,and Western blotting on days 3,7,and 14 after the injection,respectively.Moreover,the effects of COL7A1 knockdown on the proliferation and migration of hiPSCs-KCs were confirmed by immunohistochemistry,EdU,Transwell,and damage repair assays.RESULTS HiPSCs-KCs could express the hallmark proteins of KCs.COL7A1 was down-regulated in burn wound tissues and highly expressed in hiPSCs-KCs.Transplantation of hiPSCs-KCs into mice with burn wounds resulted in a significant decrease in wound area,an increase in wound re-epithelialization,a decrease in proinflammatory factors content,and an inhibition of NF-κB pathway activation compared to the PBS group.The in vitro assay showed that COL7A1 knockdown could rescue the inhibition of hiPSCs-KCs proliferation and migration,providing further evidence that COL7A1 speeds up burn wound healing by limiting cell proliferation and migration.CONCLUSION In deep,second-degree burn wounds,COL7A1 can promote KC proliferation and migration while also suppressing the inflammatory response.
基金supported by a grant from the National Natural Science Foundation of China(No.81974308).
文摘Objective Psoriasis is often combined with metabolic abnormalities,such as obesity and diabetes.The upregulation of chemerin,which is an essential protein produced primarily from white fat,is strongly correlated to the development of psoriasis.However,there is no clarification on its exact function and mechanism in disease pathogenesis.The present study aims to determine its function and mechanism in disease pathogenesis.Methods The present study used a psoriasislike inflammatory cell model and imiquimod(IMQ)-induced mouse model to confirm whether chemerin is upregulated in psoriasis patients.Results Chemerin enhanced the keratinocyte proliferation,inflammatory cytokine secretion,and activation of the MAPK signaling pathway.Crucially,the intraperitoneal injection of neutralizing anti-chemerin antibody(ChAb)diminished the epidermal proliferation and inflammation in the IMQ-induced mouse model.Conclusion The present results indicate that chemerin promotes keratinocyte proliferation,and enhances the production of inflammatory cytokines,thereby aggravating the psoriasis.Thus,chemerin can be a prospective target for the treatment of psoriasis.
文摘Background: Sugar moiety of macromolecules is today very well known for its implications in many biological recognition mechanisms including cell-cell, extracellular matrix-cell and/or bacteria-cell interactions. In this context lectins, which are carbohydrate-binding proteins displaying a high affinity for sugar groups of other molecules, are of a great importance, notably in immune response involving bacteria, viruses and fungi. As protein-carbohydrate interactions are often mediated by ions such as calcium, zinc or magnesium, we were prompted to study the effect of a thermal spring water (which contains this type of component) on interactions existing between: 1) osidic receptors of human normal keratinocytes and 2) two lectins greatly implicated in the immune response mechanisms (i.e. the dectin-1 and the langerin), and their ligands. Materials and Methods: In a first series of experiments, we studied the effect of increasing concentrations of a thermal spring water on interactions existing between glycosylated molecules and the osidic receptors expressed at the normal human keratinocytes surface. In a second step, and in order to better understand the putative effect of our thermal spring water on the immune response, we analyzed its effect on the interactions existing between the dectin-1 (implicated in the recognition of bacteria, viruses and fungi) and the langerin (expressed by Langerhans cells, the immune cells of the cutaneous tissue), and their ligands in a model using recombinant human lectins and appropriate binding molecules. Results: We showed here that our thermal spring water was able to reinforce interactions between keratinocytes osidic receptors and some of their ligands, in a dose-related manner: From 8% to 55% of increase with 10% to 30% (v/v) of thermal spring water. In the second part of our studies, we also showed that our thermal spring water was able to modulate interactions between dectin-1 and langerin and their ligands through a biphasic effect: Interactions were enhanced by more than 40% and 20% respectively with 10% of thermal spring water, and return to their basal level or lower for higher concentrations. Conclusion: The tested thermal spring water, probably due to its ionic composition, could significantly affect interactions of osidic receptors with their ligands. This property could be of a great interest to help immune system to maintain an appropriate “vigilance state” by using the thermal water at up to a concentration of 10%, and by avoiding any runaway reaction in case of aggression, by using concentrations higher than 10%. .
文摘Using inducers to induce cells to produce inflammatory response is a common in vitro experimental method to study inflammation.However,there are relatively few inflammatory models developed for the cosmetic industry,and there are also great differences in the control of model induction,the selection of inflammatory indicators,and the concentration of inducers.Therefore,in this paper,by systematically studying the effects of Lipopolysaccharide(LPS)on the cell viability,the levels of IL-1α,IL-8 and ROS of human primary keratinocytes,a skin inflammation model based human primary keratinocyte was developed.The results showed that 0.01~100μg/mL LPS had no significant effect on the cell viability of human primary keratinocytes,while 100μg/mL LPS could simultaneously induce human primary keratinocytes to produce large amounts of IL-1αand IL-8,and 0.01μg/mL LPS could induce plentiful ROS.Therefore,a skin inflammation model for differential induction of different inflammatory indicators was established,and the sample OSM2021041301 was tested with this model,it was found that sample OSM2021041301 could significantly inhibit LPS-induced elevated IL-1αand IL-8 levels,the inhibitory effect on LPS-induced elevated ROS level was weak.The results indicated that OSM2021041301 has certain anti-inflammatory effect on inflammation caused by the increase of IL-1α,IL-8 and ROS induced by LPS and its analogues.
基金Supported by the National Natural Science Foundation of China(No.81773331)CAMS Initiative for Innovative Medicine(No.2017-I2M-B&R-01)
文摘Objective Psoriasis is an immune-mediated inflammatory disease.Despite advances in the study of its pathogenesis,the exact development mechanism of psoriasis remains to be fully elucidated.Hyperproliferative epidermis plays a crucial role in psoriasis.This study aimed to investigate the effects of interleukin-36β(IL-36β)on keratinocyte dysfunction in vitro.Methods Human keratinocyte cell lines,HaCaT cells,were treated with 0(control),50 or 100 ng/ml IL-36βrespectively for 24 h.Cell viability was determined with a cell counting kit-8 assay.Flow cytometry was used to assess the effects of IL-36βon apoptosis and cell cycle distribution.Expressions of the differentiation markers,such as keratin 10 and involucrin,were evaluated by quantitative real-time polymerase chain reaction(RT-qPCR).Expressions of the inflammatory cytokines,IL-1βand IL-6 were tested by ELISA.Results CCK8 assay showed the survival rate had no significant difference between the control and treated group(P>0.05).Flow cytometry analysis showed cell cycle arrest at S phase in the IL-36β-treated groups compared with the control group(P<0.05).RT-qPCR verified the decreased mRNA expressions of keratin 10 and involucrin in the IL-36β-treated groups compared with the negative control(P<0.01).ELISA showed 100 ng/ml IL-36βenhanced levels of IL-1βand IL-6 in culture supernatants of HaCaT cells compared with the negative control(P<0.05).Conclusion Taken together,these findings suggest that IL-36βcould induce cell cycle arrest at S phase,inhibit keratin 10 and involucrin expressions and promote inflammatory activity in HaCaT cell lines.
文摘AIM:To investigate the healing process after severe corneal epithelial damage in rats treated with mesenchymal stem cells(MSCs)cultured with or without keratinocyte growth factor(KGF-2)and autologous serum(AS)on amniotic membrane(AM).Many patients are blind and devastated by severe ocular surface diseases due to limbal stem cell deficiency.Bone marrow-derived MSCs are potential sources for cellbased tissue engineering to repair or replace the corneal tissue,having the potential to differentiate to epithelial cells.METHODS:The study included 5 groups each including 10 female'Sprague Dawley'rats in addition to20 male rats used as bone marrow donors.Group I rats received AM+MSCs,Group II rats AM+MSCs cultured with KGF-2,Group III rats AM+MSCs cultured with KGF-2+AS,Group IV rats only AM and Group V rats,none.AS was derived from blood drawn from male rats and bone marrow was obtained from the femur and tibia bones of the same animals.Therapeutic effect was evaluated with clinical,histopathological and immunohistochemical assessment.MSC engraftment was demonstrated via detection of donor genotype(Y+)in the recipient tissue(X)with polymerase chain reaction.RESULTS:Corneal healing was significantly better in Groups I-III rats treated with MSC transplantation compared to Group IV and Group V rats with supportive treatment only.The best results were obtained in Group III rats with 90%transparency,70%lack of neovascularization,and 100%epithelium damage limited to less than 1/4 of cornea.CONCLUSION:We suggest that culture of MSCs with KGF-2 and AS on AM is effective in corneal repair in case of irreversible damage to limbal stem cells.
基金Supported by Postdoctoral Science Foundation of China,No.20060390192,200801243research grant from Science and Technology Department of Gansu Province,China,No.0708NKCA128
文摘AIM:To investigate the effect of keratinocyte growth factor(KGF) gene therapy in acetic acid-induced ulcerative colitis in rat model.METHODS:The colitis of Sprague-Dawley rats was induced by intrarectal infusion of 1 mL 5%(v/v) acetic acid.Twenty-four hours after exposed to acetic acid,rats were divided into three experimental groups:control group,attenuated Salmonella typhimurium Ty21a strain(SP) group and SP strain carrying human KGF gene(SPK) group,and they were separately administered orally with 10% NaHCO3,SP or SPK.Animals were sacrificed and colonic tissues were harvested respectively on day 3,5,7 and 10 after administration.Weights of rats,colonic weight/length ratio and stool score were evaluated.Histological changes of colonic tissues were examined by hematoxylin and eosin(HE) staining method.The expression of KGF,KGF receptor(KGFR) and TNF-α were measured either by enzyme-linked immunosorbent assay or Western blotting.Immunohistochemistry was used to detect the cellular localization of KGFR and Ki67.In addition,superoxide dismutase(SOD) activity and malondialdehyde(MDA) contents in the homogenate were measured.RESULTS:Body weight and colonic weight/length ratio were declined in SPK group compared with SP and control groups(body weight:272.78 ± 17.92 g vs 243.72 ± 14.02 g and 240.68 ± 12.63 g,P < 0.01;colonic weight/length ratio:115.76 ± 7.47 vs 150.32 ± 5.99 and 153.67 ± 5.50 mg/cm,P < 0.01).Moreover,pathological changes of damaged colon were improved in SPK group as well.After administration of SPK strain,KGF expression increased markedly from the 3rd d,and remained at a high level till the 10th d.Furthermore,KGFR expression and Ki67 expression elevated,whereas TNF-α expression was inhibited in SPK group.In the group administered with SPK,SOD activity increased significantly(d 5:26.18 ± 5.84 vs 18.12 ± 3.30 and 18.79 ± 4.74 U/mg,P < 0.01;d 7:35.48 ± 3.35 vs 22.57 ± 3.44 and 21.69 ± 3.94 U/mg,P < 0.01;d 10:46.10 ± 6.23 vs 25.35 ± 4.76 and 27.82 ± 6.42 U/mg,P < 0.01) and MDA contents decreased accordingly(d 7:7.40 ± 0.88 vs 9.81 ± 1.21 and 10.45 ± 1.40 nmol/mg,P < 0.01;d 10:4.36 ± 0.62 vs 8.41 ± 0.92 and 8.71 ± 1.27 nmol/mg,P < 0.01),compared with SP and control groups.CONCLUSION:KGF gene therapy mediated by attenuated Salmonella ameliorates ulcerative colitis induced by acetic acids,and it may be a safe and effective treatment for ulcerative colitis.
文摘Objective:To examine the level of expression of anti skin aging gene of Ganoderma lucidum polysacchayides and clarify its mechanism with anti aging of this ancient Chinese medicine.Methods:HacaT cell of keratinocytes lines were cultured and treated with the polysaccharides.The total RNA was extracted with Trizol reagent and cDNA was synthesized by reverse thanscription.The obtained cDNAs were then fluorescently labeled with cy3 and cy5 respectively and hybridized with gene expressing pedigree cDNA chip.The images were scanned and analyzed with special software.The scan data were analyzed with software and checked by real time PCR.Results:Among total 18 346 human genes,the expression of 103 ones was up-regulated and 378 ones down-regulated.It was demonstrated evidently that Ganoderma lucidum polysaccharides affected the expression of genes of anti skin aging.Two ways are anastomotic.Conclusion:it is concluded by analysis of function of these up-regulation and down-regulation genes that Ganoderma lucidum polysaccharides may play an important role in boosting cell growth and against skin aging.It shows that the results of gene array reliable by real time PCR.
文摘Objective:To investigate the effect of lead on the production of IL-8 and cell proliferation in normal human oral keratinocytes(NHKs).Methods:NHKs were prepared as outgrowths from normal human buccal mucosa.The cells were treated with three concentrations of lead glutamate(4.5×10<sup>-5</sup>M,4.5×10<sup>-6</sup>M and 4.5×10<sup>-7</sup>M).NHKs grown in glutamic acid were used as control.The amounts of IL-8 secreted in the culture supernatants were evaluated at 12 and 24 h using enzyme-linked immunospecific assay(ELISA).Cell proliferation was determined by the MTT colorimetric assay.Three cultures were used for each experiment,and three independent experiments were performed.Analysis of variance and Duncan’s multiple range tests were used for statistical analysis.Results:An elevation of IL-8 in culture supernatants of NHKs treated with lead at all concentrations at 12 and 24 h after exposure in a dose-dependent manner was revealed.A significant increase in cell numbers was observed only at 24 h exposed to 4.5×10<sup>- 5</sup>M lead glutamate.Conclusions:The capacity of NHKs,to secrete IL-8,enhanced by lead glutamate,is demonstrated here.Induction of cell proliferation is revealed only after exposure to high lead concentration.The elevation of secreted IL-8 is a probable initial sign for the acute inflammatory response and may be involved in the pathogenesis of lead stomatitis.
基金supported by grants from the National Natural Science Foundation of China(No.81171495,No.81271765,and No.81400970)
文摘Liopxin A4(LXA4) is considered to be a crucial modulator in the inflammatory responses. In the present study, we aimed to study the effect of LXA4 on the inflammatory cytokines production induced by lipopolysaccharide(LPS) and the possible mechanism in normal human epidermal keratinocytes(NHEKs). NHEKs were isolated and cultured. The expression of toll-like receptor 4(TLR4), LXA4 receptor(ALXR) and aryl hydrocarbon receptor(Ah R) in NHEKs was detected by reverse transcription polymerase chain reaction(RT-PCR). The m RNA and protein levels of tumor necrosis factor-alpha(TNF-α) and interleukin-1β(IL-1β) were determined in NHEKs stimulated by LPS(10 μg/m L) with or without preincubation with LXA4(100 nmol/L) for 30 min by real-time quantitative PCR(real-time q PCR) and enzyme-linked immunosorbent assay(ELISA), respectively. The expression levels of tumor necrosis factor receptor-associated factor 6(TRAF6) and suppressors of cytokine signaling 2(SOCS2) m RNAs and proteins, and nuclear translocation of NF-k B-p65 were measured by real-time q PCR and Western blotting, respectively. The results showed that NHEKs expressed TLR4, ALXR and Ah R. LXA4 significantly inhibited the m RNA and protein expression levels of TNF-α, IL-1β and TRAF6 induced by LPS in NHEKs, and LXA4 obviously increased the expression of SOCS2 at m RNA and protein levels. The nuclear NF-k B-p65 protein expression induced by LPS was inhibited after preincubation with LXA4 in NHEKs. It was concluded that LXA4 inhibits the LPS-induced production of TNF-α and IL-1β in NHEKs by up-regulating SOCS2 and down-regulating TRAF6.
基金supported by National Natural Science Foundation of China(No.81441126)
文摘The most effective sequence of small interfering RNA(si RNA) silencing STAT3 of psoriatic keratinocytes(KCs) was screened out,and the effects of the most effective si RNA combined with ultrasonic irradiation and Sono Vue microbubbles on the expression of STAT3 of KCs and the dose-and time-response were investigated.Three chemically-synthetic si RNAs targeting STAT3 carried by Lipofectamine 3000 were transfected into KCs,and the effects on STAT3 expression were detected,then the most effective si RNA was selected for the subsequent experiments.The negative controls of siR NA(si RNA-NC) labeled with Cy3 carried by Lipofectamine 3000 combined with ultrasonic irradiation and Sono Vue microbubbles were transfected into KCs,then the optimal parameters of ultrasonic irradiation were determined.The most effective si RNA carried by Lipofectamine 3000 combined with ultrasonic irradiation at the optimal parameters and Sono Vue microbubbles was transfected into KCs,and the dose-and time-response of RNA interference was determined.The effect of RNA interference by the most effective si RNA at the optimal time and dose carried by Lipofectamine 3000 combined with ultrasonic irradiation and Sono Vue microbubbles(LUS group) was compared with that only carried by Lipofectamine 3000(L group).The results showed that si RNA-3 achieved the highest silencing efficacy.0.5 W/cm2 and 30 s were selected as the parameters of ultrasonic irradiation.The si RNA-3 carried by Lipofectamine 3000 combined with ultrasonic irradiation and Sono Vue microbubbles could effectively knock down the STAT3 expression at m RNA and protein levels in dose-and time-dependent manners determined at 100 nmol/L with maximum downregulation on m RNA at 48 h,and on protein at 72 h after transfection.The LUS group achieved the highest silencing efficacy.It was concluded that si RNA-3 carried by Lipofectamine 3000 combined with ultrasonic irradiation and SonoV ue microbubbles could effectively knock down the STAT3 expression in psoriatic KCs,and the optimized transfection condition and the sequence of si RNA-3 could serve for further research on gene therapy of psoriasis.