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Background: Generalized vitiligo is a common, multifaceted, polygenic condition in which autoimmune loss of melanocytes results in depigmented skin patches, overlying hair and mucous membranes. NLRP1 has been proposed to be implicated in the susceptibility of a broad variety of autoimmune disorders, including generalized vitiligo (GV). Genetic polymorphisms in the NLRP1 encoding gene (formerly known as NALP1) have previously been found to be linked with GV and there is uncertainty as to their role in the modulation of NLRP1 expression. Oxidative stress is a significant pathogenesis theory for vitiligo. Glutathione S-transferases (GSTs) are enzymes active in the defense of cells against chemical toxicity and stress.This study validates some of the Unani concepts of humors or temperaments (Phenotypes), with regard to Vitiligo, Where vitiligo is regarded as a phlegmatic disease. We selected Vitiligo subjects with Phlegmatic Clinical Phenotype for our study, with an aim to determine its association with the genetic biomarkers- NLRP1, GSTM1 and GSTT1 null genotypes and other biochemical parameters. Methods: The Unani clinicians randomly selected 100 vitiligo patients with a phlegmatic Clinical Phenotype who were attending NRIUMSD for treatment and 100 healthy volunteers belonging to Phlegmatic (Phlegmatic clinical Phenotype). Besides looking at temperaments/ humors as susceptibility factors – we included a genetic factor- NLRP1, GSTM1- and GSTT1-null genotypes to our investigation. We have genotyped the NLRP1, GSTM1- and GSTT1-null genotypes by PCR-RFLP and by Multiplex PCR, GST protein level estimation by ELISA method. Results: NLRP1 rs2670660 polymorphism was shown to be in significant association with GV, with the presence of minor alleles in active GV. We found that the frequencies of GSTM1 null genotype and GSTT1 null genotype in vitiligo patients were significantly high compared to the controls (OR= 1.47, 95% CI=0.765--2.861), (OR = 4.75, 95% CI = 2.131-10.63), respectively. In combination analysis with both genes, the results suggested significant association of vitiligo risk with both GSTM1\GSTT1 null genotypes (OR=4.83, 95% CI=1.523– 15.32).We observed a significant decrease (p<0.001) in GST protein levels. Conclusion: Our findings indicate that NLRP1 rs2670660 polymorphism may be genetic risk factor for susceptibility to GV and the null genotypes of GSTM1 and GSTT1 of both genes increase the risk of the disease. A significant decrease (p<0.001) in GST protein levels appeared to be a key feature in Vitiligo subjects, Therefore, detection of antioxidant enzyme levels can be effective biomarkers for early detection of the disease. We believed that GSTM1-and GSTT1-null genotype polymorphisms were associated with an increased risk of vitiligo. This is the first study of its kind along with Clinical Phenotype as per Unani Philosophy.

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Phenotypic heterogeneity is one of the major concerns in personalized medicine. Variations in drug metabolism rates vary in every individual and have been associated with drug metabolizing gene polymorphism. CYP3A4*2 gene polymorphism in drug metabolism has been reported to be associated with a range of diseases. In this regard, the present investigation is aimed to determine the possible association of Unani philosophy of temperaments (clinical phenotypes) with CYP3A4*2 gene polymorphism in patients and controls. In the present study, 800 subjects were analyzed for CYP3A4*2 gene polymorphism by PCR-RFLP technique in four different clinical phenotypes (Sanguine, Phlegmatic, Choleric and Melancholic) as per unani philosophy. Mutant allelic frequencies of patients and controls of four unani temperaments are sanguine (Damavi) (0.19 vs 0.12), phlegmatic (Balghami) (0.21 vs 0.11), bilious (Safravi) (0.23 vs 0.11) and melancholic (Saudavi) (0.16 vs 0.11) subjects. Safravi (p<0.04) and Balghami (p<0.01) results are statistically significant. Bilious (Safravi) and phlegmatic (Balghami) patients showed significant association with CYP3A4*2 gene polymorphism with 1.76 and 2.04 odds risk. Further such studies are essential in unani regime for providing better treatment for many diseases which require personalized medicine based on phenotype-genotype correlation.

In view inflammation causes of development of some chronic diseases like rheumatoid arthritis, obesity, diabetes, asthma. Cell model in vitro in an effort to provide an understanding about the cellular and molecular mechanism of Unani formulation are being used for treatment of various inflammatory diseases including arthritis for centuries. Method Antioxidant by DPPH assay. Cell viability through MTT assay. Measurement of NO level and ROS levels by (DCFDA) in RAW cells. Results in DPPH Assay showed all the extracts ofUNIM-302 shows good antioxidant activity when compared to UNIM301. UNIM drugs show no substantive cytotoxic activity against RAW macrophages cell line. UNIM301, UNIM 302 inhibited the production of Nitric oxide production in RAW cells. ROS the results found that UNIM301 and UNIM302 of all extracts could significantly inhibits the LPS stimulation and reduces production of ROS in RAW cells. In conclusion study demonstrated that both Unani formulations inhibits NO and ROS production in LPS induced RAW macrophages. Activity in all different extracts UNIM 302 exhibited better antioxidant compared to UNIM 301these unim formulation extracts can be used as natural sources of antioxidants potent anti inflammatory agent and exhibits inflammatory preventive properties.