Chemical chaperone, TUDCA unlike PBA, mitigates protein aggregation efficiently and resists ER and non-ER stress induced HepG2 cell death.

Stress induced BSA (bovine serum albumin) protein aggregation is effectively mitigated in vitro by TUDCA (tauroursodeoxycholic acid) than by PBA (4- phenylbutyric acid), chemical chaperones approved by FDA for the treatment of biliary cirrhosis and urea cycle disorders respectively. TUDCA, unlike PBA, enhances trypsin mediated digestion of BSA. TUDCA activates PERK, an ER-resident kinase that phosphorylates the alpha-subunit of eukaryotic initiation factor2 (eIF2α) and promotes the expression of activated transcription factor 4 (ATF4) in HepG2 cells. In contrast, PBA induced eIF2α phosphorylation is not mediated by PERK activation and results in low ATF4 expression. Neither chaperones promote expression of BiP, an ER chaperone, and CHOP (C/EBP homologous protein), downstream target of eIF2α-ATF4 pathway. Both chaperones mitigate tunicamycin induced PERK-eIF2α-ATF4-CHOP arm of UPR and expression of BiP. TUDCA, unlike PBA does not decrease cell viability and it also mitigates tunicamycin, UV-irradiation and PBA induced PARP (poly ADP-ribose polymerase) cleavage and cell death. These findings therefore suggest that TUDCA's antiapoptotic activity to protect HepG2 cells and PBA's activity that limits tumor cell progression may be important while considering their therapeutic potential.

Tauroursodeoxycholic acid prevents stress induced aggregation of proteins in vitro and promotes PERK activation in HepG2 cells.

Tauroursodeoxycholic acid (TUDCA) a bile salt and chemical chaperone reduces stress-induced aggregation of proteins; activates PERK [PKR (RNA-dependent protein kinase)-like ER (endoplasmic reticulum) kinase] or EIF2AK3, one of the hall marks of ER stress induced unfolded protein response (UPR) in human hepatoblastoma HepG2 cells; prevents heat and dithiothreitol (DTT) induced aggregation of BSA (bovine serum albumin), and reduces ANS (1-anilino-naphthalene-8-sulfonate) bound BSA fluorescence in vitro. TUDCA inactivates heat treated, but not the native EcoR1 enzyme, and reduces heat-induced aggregation and activity of COX-1 (cyclooxygenase enzyme-1) in vitro. These findings suggest that TUDCA binds to the hydrophobic regions of proteins and prevents their subsequent aggregation. This may stabilize unfolded proteins that can mount UPR or facilitate their degradation through cellular degradation pathways.

Role of oxidative stress and autoimmunity in onset and progression of vitiligo.

Vitiligo is an acquired depigmentation disorder characterized by the loss of functional melanocytes from the epidermis. Two major theories of vitiligo pathogenesis include autoimmunity and oxidative stress-mediated toxicity in melanocytes. The present study aimed to evaluate both the hypotheses in vitiligo patients and to investigate their role in the disease onset and progression. Antimelanocyte antibody levels and lipid peroxidation (LPO) levels were evaluated in 427 patients and 440 controls; antithyroid peroxidase (TPO) antibody levels were estimated in 102 patients and 72 controls. Patients showed a significant increase in LPO and antimelanocyte antibody levels compared to controls. Antimelanocyte antibody and LPO levels were higher in active vitiligo compared to stable. Only 9.8% of patients showed the presence of anti-TPO antibodies in their circulation. Oxidative stress may be the initial triggering event to precipitate vitiligo in Gujarat population, which is exacerbated by contributing autoimmune factors together with oxidative stress.

Tumor necrosis factor B (TNFB) genetic variants and its increased expression are associated with vitiligo susceptibility.

Genetic polymorphisms in TNFB are involved in the regulation of its expression and are found to be associated with various autoimmune diseases. The aim of the present study was to determine whether TNFB +252A/G (rs909253) and exon 3 C/A (rs1041981) polymorphisms are associated with vitiligo susceptibility, and expression of TNFB and ICAM1 affects the disease onset and progression. We have earlier reported the role of TNFA in autoimmune pathogenesis of vitiligo, and we now show the involvement of TNFB in vitiligo pathogenesis. The two polymorphisms investigated in the TNFB were in strong linkage disequilibrium and significantly associated with vitiligo. TNFB and ICAM1 transcripts were significantly increased in patients compared to controls. Active vitiligo patients showed significant increase in TNFB transcripts compared to stable vitiligo. The genotype-phenotype analysis revealed that TNFB expression levels were higher in patients with GG and AA genotypes as compared to controls. Patients with the early age of onset and female patients showed higher TNFB and ICAM1 expression. Overall, our findings suggest that the increased TNFB transcript levels in vitiligo patients could result, at least in part, from variations at the genetic level which in turn leads to increased ICAM1 expression. For the first time, we show that TNFB +252A/G and exon 3 C/A polymorphisms are associated with vitiligo susceptibility and influence the TNFB and ICAM1 expression. Moreover, the study also emphasizes influence of TNFB and ICAM1 on the disease progression, onset and gender bias for developing vitiligo.

Involvement of superoxide dismutase isoenzymes and their genetic variants in progression of and higher susceptibility to vitiligo.

Oxidative stress has been implicated as the initial triggering event in vitiligo pathogenesis leading to melanocyte destruction. Here, we report a significant increase in oxidative stress in vitiligo patients as evidenced by high lipid peroxidation levels suggesting an imbalance in the antioxidant enzyme system as reported in our previous studies. This study examined the role of the enzymatic antioxidant SOD, which converts the pro-oxidant superoxide into H2O2, in vitiligo pathogenesis. The activity of three isoforms of SOD, i.e., SOD1, SOD2, and SOD3, was significantly higher in vitiligo patients. To identify the underlying mechanism for the increase in activities of SOD isoforms, we explored the SOD1, SOD2, and SOD3 genes for their genetic variations and transcript levels. The SOD2 Thr58Ile (rs35289490) and Leu84Phe (rs11575993) polymorphisms were significantly associated with vitiligo patients, and the Val16Ala (rs4880) polymorphism was associated with active vitiligo patients. Interestingly, SOD2 activity was contributed by these polymorphisms along with its increase in transcript levels in patients. SOD3 activity was associated with the Arg213Gly (rs8192291) polymorphism. The SOD3 transcript levels were also increased in patients, which might contribute to the increased SOD3 activity. However, we could not establish the genotype-phenotype correlation for SOD1 as we could not detect any novel or reported SNPs in SOD1. In addition, both transcript and protein levels of SOD1 were unchanged between patients and controls, though SOD1 activity was increased in patients. Activities of SOD isoforms also correlated with progression of the disease as the activity was higher in active cases of vitiligo compared to stable cases. Here, we report that SOD2 and SOD3 polymorphisms may be genetic risk factors for susceptibility and progression of vitiligo and hence the genetic makeup of an individual may form a basis for the effective treatment of the disease. Overall, our results suggest that increased activity of SOD isoforms under the influence of genetic factors may lead to accumulation of H2O2 in cytoplasmic, mitochondrial, and extracellular compartments resulting in oxidative damage to the melanocytes.

Vitiligo: interplay between oxidative stress and immune system.

Vitiligo is a multifactorial polygenic disorder with a complex pathogenesis, linked with both genetic and non-genetic factors. The precise modus operandi for vitiligo pathogenesis has remained elusive. Theories regarding loss of melanocytes are based on autoimmune, cytotoxic, oxidant-antioxidant and neural mechanisms. Reactive oxygen species (ROS) in excess have been documented in active vitiligo skin. Numerous proteins in addition to tyrosinase are affected. It is possible that oxidative stress is one among the main principal causes of vitiligo. However, there also exists ample evidence for altered immunological processes in vitiligo, particularly in chronic and progressive conditions. Both innate and adaptive arms of the immune system appear to be involved as a primary event or as a secondary promotive consequence. There is speculation on the interplay, if any, between ROS and the immune system in the pathogenesis of vitiligo. The article focuses on the scientific evidences linking oxidative stress and immune system to vitiligo pathogenesis giving credence to a convergent terminal pathway of oxidative stress-autoimmunity-mediated melanocyte loss.