The Mechanisms Of Abnormality Of Langerhans Cells In The Lesions Of Condyloma Acuminatum

PartⅠAnalyses of immunophenotype of Langerhans cells in lesions of condyloma accuminatumBackground The density and number of LC maintain equilibrium in normal condition,the density of which is normally between 200/mm~2 to 970/mm~2,while it increases in infection and inflammation situation.CA is a kind of dermal benign tumor caused by HPV infection,there is no consensus to the character of LC in lesions of CA at present.Objective To explore the change of number,density,morphology and function of LC in lesions of CA.Methods The expression of surface markers,namely LC:CD1a, S-100 protein and HLA-DR in 30 cases of CA,were analyzed by immunohistochemistry technique,and 20 cases of normal acrobystia were taken as controls.The difference of the morphology,number and function of LC between two groups was investigated.Results1.Cell counting of LC:①There was a well-distribution of CD1a+LC in normal skin,it mainly located in cutaneous stratum spinosum and stratum basale,lower expression was found in superficial dermis.LCs were in round,oval or irregular shapes,they had slender,multi-cell and sub-cell cytodendrite.CD1a+LC in the lesions of CA scattered evenly in the stratum spinosum and stratum basale,most were seen in middle epidermis and less were seen in papillary layer of dermis.CD1a+LC were irregular,round or oval shaped,they had slender,multi-cell and sub-cell cytodendrite.The density of CD1a+LC in normal epidermis was (201.71±53.69)/mm~2 and(55.91±29.64)/1000KC,respectively,and that in lesions of CA was(195.23±56.38)/mm~2 and(52.63±31.29)/1000KC, respectively.There was no significant statistic difference between two groups(P＞0.05).②Cell counting of S-100+LC:S-100+LC mainly located above stratum basale epidermis and lower expression was found in papillary layer of dermis.LCs were round,oval or punctiform shaped,with a little short cell process.No S-100+LC was detected in some part of epidermis in some samples.There was no typical dendritic cells in lesions of CA, and only a little punctiformed cytodendrite was found.The density of S-100+LC in normal epidermis was(62.35±33.97)/mm~2 and(14.25±5.74)/ 1000KC,respectively,and that in lesions of CA was(16.12±10.84)/mm~2 and(4.06±2.68)/1000KC,respectively.There was a significant statistic difference between two groups(P＜0.05).③Cell counting of HLA-DR+LC:There was a well-distribution of CD1a+LC in normal skin,which predominantly located in epidermis acanthocyte and stratum basale,and some dendritic cells were found in superficial lamella corium.The morphology of LC was similar to that of CD1a+LC,the presence of HLA-DR antigen in normal epidermal keratinocytes was scarce.There was a little HLA-DR+LC in lesions of CA,much were seen in the middle and inferior part of epidermis,and the typical dendritic cells were decreased,the shrink or loss of LC and its cytodendrite were hardly detected.No HLA-DR antigen was found in KC of CA.The number of HLA-DR+LC in normal epidermis was (183.66±42.22)/mm~2 and(50.91±32.57)/1000KC,respectively,and that in lesions of CA was(125.26±25.13)/mm~2 and(32.38±13.80)/1000KC, respectively.There was a significant statistic difference between two groups(P＜0.05)2.In the semiquantitative analysis of IHC:20 normal skins were stained for CD1a,2 were negative stained,5 weakly positive stained, 10 positive stained and 3 strongly positive stained;and the IHC of CD1a was done in lesions of CA from 30 patients,3 were negatively stained,6 weakly positive stained,16 positive and 5 strongly positive stained,there was no significant statistic difference between two groups(P＞0.05).IHC analysis of S-100 protein was done in 20 normal skins,3 were negatively stained,14 were weakly positive and 3 were positive stained; and in the lesions of CA from 30 patients stained for S-100 protein,16 were negatively stained and 15 were weakly positive stained,there was a significant difference between two groups(P＜0.05).IHC analysis was done in 20 normal skins,3 were negative stained,6 weakly positive stained,9 positive and 2 strongly positive stained;when lesions of CA from 30 patients were stained for HLA-DR,8 negative stained,16 weakly positive and 6 positive stained were found,there was a significant statistic difference between two groups(P＜0.05).3.In the quantitative analysis of IHC,there was no significant difference in gray density of CD1a between normal control and CA group (P＞0.05).However,significant differences in the density of gray color were found in protein S-100 and HLA-DR between normal and CA group(P＜0.05),the expression of S-100+LC and HLA-DR+LC in CA were much lower than in normal control.4.The mean density of S-100+LC in epidermis of CA from 30 patients was 15.25,and the number of CD1a+LC was 195.23,the ratio of S-100+LC to CD1a+LC was 7.30%;and in control,it was 30.91%,there was a significant difference between the ratios of two groups(P＜0.05).5.To compare the mean gray scale among CD1a+LC,S100+LC and HLA-DR+LC in lesions of CA,SPSS12.0 software was used by Pearson Correlation.The linerar correlation coefficient between CD1a+LC and HLA-DR+LC in epidermis of CA was 0.5728(P＜0.05),it was demonstrated that there was a linear relativity between them;and that of CD1a+LC and S-100+LC was 0.0961(P＞0.05),there was no linear relationship between the two values;and the correlation coefficient between S-100+LC and HLA-DR+LC was 0.0392(P＞0.05),there was no linear relationship between the values as well.Conclusion The number and morphology of LC in lesions of CA were seemingly normal,but its functional maturation was inhibited,90% of LC in lesions of CA was immature,the mechanism of its functional dysmaturity is not clear.PartⅡExpression of MCP-1 in lesions of condyloma accuminatumBackground Peripheral blood mononuclear cell is the precursor cell of LC,chemotatic factor which is mainly generated from malpighian cells in the lesions is able to affect the migration of immunocyte such as monocyte.Monocyte chemoattractant protein-1(MCP-1)is one of the major chemotatic factors,but it's unclear whether the abnormality of LC in CA is induced by MCP-1 which is generated from keratinocytes in the lesions.Objective To investigate the expression of MCP-1 in the lesions of CA at protein and nucleic acid level,and to determine the effect of MCP-1 on the abnormality of LC in the lesions of CA.Methods The expression of MCP-1 protein and mRNA in 30 CA skins and 20 normal skins were analyzed by immunohistochemistry, Western Blotting and hybridization in situ.Results1.In the in situ hybridization analysis,the yellow granulo-like MCP-1mRNA staining was occasionally found either in endochylema or cellular nucleus.Weak staining of MCP-1mRNA was found in endochylema of epiderm keratinocytes in normal skin,especially in the stratum basale,whereas there was no staining of MCP-1mRNA in CA. There was no significant difference compared to normal skin group(P＞0.05);2.In immunohistochemistry analysis,the yellow granulo-like MCP-1 protein was occasionally seen in either endochylema of keratinocytes or cellular membrane,weakly positive staining of MCP-1 was found in the stratum basale of normal skin,and in the tissues of sebaceous gland and coil gland.MCP-1 protein was either undetectable or expressed at extremely low level in condyloma accuminatum.There was no significant difference as compared to that in normal skins(P＞0.05);3.In Western Blotting analysis,the expression of MCP-1 in CA and normal skins were extremely low,the statistic difference is not significant (P＞0.05)Conclusion MCP-1 was expressed at low level in keratinocytes of CA,the small amount of MCP-1 was not capable of inducing the monocyte transformation to LC and hence the inflammatory and immune reaction will not be staged. PartⅢExpression of haptoglobin in lesions of condyloma accuminatumBackground There were no obvious abnormality of morphology and quantity of LC in the local lesions of CA,but S-100+DC were rarely found.The results suggest that there was a disturbance of functional maturation of LC,which may be caused by the abnormality of some cytokines that they are not able to induce LC to be mature despite of the stimulating by HPV antigen.We used haptoglobin(Hp)which can inhibit the functional maturation of LC to study the mechanism of abnormality of LC in CA.Objective To investigate the expression of haptoglobin in lesions of condyloma acuminatum at mRNA and protein level;and explore its role in the pathogenesis of CA.Methods The expression of Hp protein and mRNA in skin tissues from 30 patients with CA and from 20 normal controls were detected by immunohistochemistry(IHC),Western Blot and hybridization in situ.Results1.In situ hybridization study showed that the weakly positive signals of Hp mRNA were detected in the endochylema of keratinocytes,which were found predominantly in the stratum basale and stratum spinosum. Hp mRNA was expressed in the keratinocytes in lesions of CA,the distribution of Hp mRNA expression in lesions of CA was similar as compared with normal controls,but higher expression of Hp mRNA was found in CA than in nomal controls,There was a significant difference in the positive expression of HpmRNA between CA group and control group(P＜0.05);2.Immunohistochemical study showed that Hp protein was expressed in the keratinocytes in lesions of CA at high level,Hp protein was expressed in the whole layers of epidermal keratinocytes in the lesions of CA,stronger staining was seen in the stratum basale and stratum spinosum.Hp protein was expressed predominantly in the stratum basale in normal skin,while weak staining was seen below the stratum spinosum.The positive ratio of Hp expression was 93.3%and 80.0%in CA group and normal controls,respectively.There was a significant statistic difference between two groups(P＜0.05).There was a significant difference in the mean density of gray color between CA group and control group(P＜0.05);3.Western Blot showed that the expression of Hp in CA lesions was significantly increased compared with normal skins(P＜0.05)Conclusion The expression of Hp mRNA in keratinocytes in the lesions in CA was significantly increased,in addition the keratinocytes in the lesions of CA were able to synthesize Hp protein.Hp in lesions of CA participated in the local immunity escape by which Hp might prevent LC functional maturation and might inhibit the immunocompetence of KC.