Study On The Expressions And Their Significance Of Aryl Hydrocarbon Receptor And Related Genes In Human Sebaceous Gland Cells

With the rapid development of modern industry has created great benefits forhumanity, but all too often has come at the price of escalating health hazards arisingfrom unwanted industrial by-products and chemical waste. As an important ofby-products of industrial processes, dioxins has been known as the representative of aseries of environmental pollutants. There are about75different dioxin isomers, perhapsthe most vivid example of toxic chemical agents is the halogenated hydrocarbon2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) that serves as the prototype for a largegroup of polyhalogenated dibenzo-para-dioxins which evoke a wide range of adversetoxic effects in human and animals. TCDD is associated with developmental toxicity,carcinogenicity, immunotoxicity, endocrine toxicity and skin toxicity, which havelong-term effects on human beings’ health. Thus, understanding the impact of TCDD onhuman health has became a research focus.Chloracne, a hyperkeratotic skin disorder with acne-like eruption of comedonesand cysts of pilosebaceous unit in facial sebaceous glands, is the most consistentmanifestation of dioxin intoxication and is considered to be its reliable hallmark, whichmainly displays the comedo and cysts The important features of this disease contain the atrophy of sebaceous glands and sebum hyposecretion. Histopathology exhibits atrophyor absence of sebaceous glands as well as infundibular dilatation or cystic formation ofhair follicles, hyperplasia of epidermis, and hyperpigmentation of stratum corneum.Therefore, it is great significant for revealing the mechanisms of chlorance to charifythe relationship between dioxin and sebaceous gland differentiaion, as well as the lipidsynthesis. Since previous studies have proved that TCDD effects human sebocytes weremediated through the aryl hydrocatbon receptor(AhR) signaling pathway, and inducedthe sebaceous switching into keratinocyte differentiation in vitro, which was proposedto be the important mechanism of chloacne. So based on this, we further study thesignaling pathway of TCDD induced sebocytes abnormal differentiation.The current researches have showed that TCDD functions mostly via thearylhydrocarbon receptor (AhR) signaling pathway. As the ligand of AhR, Ah receptornuclear translocator protein (ARNT) is involved in the formation of the AhR/ARNTheterodimer complex which then is capable of activating the downstream target genes.Cytochrome P4501A1(CYP1A1) is identified as the downstream target gene oncanonical AhR signaling transduction in many cells. In recent years, Blymphocyte-induced maturation protein1(Blimp1) has also been concerned as anothernew target gene on the non-canonical AhR signaling pathway. However, which pathwayTCDD gets through to mediate the differentiation abnormality of sebaceous gland cellsis still unknown. And how TCDD activates the downstream target genes of cellularsignaling pathway is also unclear. Therefore, further studies are needed to solve theabove issues. Therefore, We hypothesized that TCDD-induce the differentiationabnormality of human sebocytes may be via AhR/ARNT signaling pathway, meanwhileCYP1A1and Blimp1probably play an important role on the molecular pathogenesis ofchloracne as downstream target genes.Objectives:An culture model in vitro was developed, taking the immortalized SZ95sebocytes and sebaceous glands from human skin tissues as samples, to explore the mechanisms ofTCDD signaling on cytotoxicity and investigate the downstream target genes activatedby TCDD, which can reveal the TCDD signaling on human sebocytes and provideevidence to explain the molecular mechanism of chloracne.Methods:①Immunocytochemistry method was used to study on AhR、ARNT、CYP1A1and Blimp1proteins expression in SZ95cells, and used to study the effects of10nMTCDD for3days on the above gene protein expressions.②Western blotting was used to study on AhR、ARNT、CYP1A1and Blimp1proteins expression in SZ95cells., and used to study the effects of10nM TCDD for3days on the above gene protein expressions.③Immunohistochemistry method was used to study on AhR、ARNT、CYP1A1and Blimp1proteins expression in human facial sebaceous gland tissues, and used tostudy the effects of10nM TCDD for3days on the above gene protein expressions.④HE staining was applied for detecting the changes of sebaceous gland tissuesfrom human facial skin7days after treated with10nM TCDD.Results:①Immunocytochemistry (ICC) method results showed that AhR and CYP1A1were localized mainly in the cytoplasm of SZ95human sebocytes but the ARNT andBlimp1were mainly present in the nucleus. After treated with10nM TCDD for3days,the expression of AhR significantly decreased while the expression of ARNT、CYP1A1and Blimp1were up-regulated in SZ95sebocytes.②Western blot results showed that the expression of AhR proteins in SZ95sebocytes treated by10nM TCDD for3days was lower than that in negative controlgroup(0.153±0.025vs.0.343±0.021, P<0.05); the expression of ARNT proteins treatedby10nM TCDD for3days was higher than that in negative control group(2.090±0.362 vs.0.997±0.156, P<0.05); the expression of CYP1A1proteins treated by10nM TCDDfor3days was significantly higher than that in negative control group(4.233±0.252vs.0.123±0.208, P<0.05); the expression of Blimp1proteins treated by10nM TCDD for3days was significantly higher than that in negative control group(2.930±0.058vs.0.440±0.036, P<0.05).③Immunohistochemistry(IHC) method results showed that AhR、ARNT、CYP1A1and Blimp1proteins were expressed in sebaceous gland tissue from humanfacial skin. After treated with10nM TCDD for3days, the expression of AhRsignificantly decreased while the expression of ARNT、CYP1A1and Blimp1wereup-regulated in sebaceous gland tissue.④HE staining results showed that some keratinocyte-like cells emerged around thehuman sebaceous gland tissue7days after treated with10nM TCDD, accompanyingwith the atrophy of sebaceous glands.Conclusions:Our study found the expression of AhR, ARNT, CYP1A1and Blimp1proteins inSZ95human sebocytes as well as in sebaceous glands from human scalp and facial skintissues, which can be activated by TCDD. The experiment in vitro implied thatTCDD-induce the differentiation abnormality of human sebocytes may be viaAhR/ARNT signaling pathway. CYP1A1and Blimp1probably play an important roleon the molecular pathogenesis of chloracne as downstream target genes.