“TO COMPARE BETWEEN DIRECT IMMUNOFLUORESCENCE OF SKIN BIOPSY & PLUCKED HAIR FOLLICLES IN PATIENTS WITH PEMPHIGUS”.
BRIEF RESUME OF THE INTENDED WORK:
6.1 Need for study:
Pemphigus is a autoimmune blistering disease presenting clinically with flaccid intraepithelial blisters, erosions, and ulcerations of the skin and mucous membranes.In these dermatosis antibodies are formed against the antigens involved in adhesion between epidermal cells.These antibodies are demonstrable by direct immunofluorescence (DIF) testing of skin, and indirect immunofluorescence (IIF) testing of serum.
Direct immunofluorescence examination of the perilesional uninvolved skin will show the presence of antibodies of IgG,IgM,IgA class & the complement C3 component between epidermal cells. This method is the gold standard in the diagnosis of pemphigus.
A specific immunofluorescence pattern is demonstrated in the outer root sheath of anagen and telogen hair.. Hence the study is to analyse the direct immunofluorescence (DIF) of skin biopsy & plucked hair follicles in patients with pemphigus.
.6.2 REVIEW OF LITERATURE:
A study by Rao R, et al have shown that Intercellular deposition of IgG was seen in the outer root sheath of anagen hair in 85% of cases. They have concluded Direct immunofluorescence (DIF) of hair follicles is a simple, non-invasive test
Kumaresan, Reena Rai, and V Sandhyaet al performed direct immunofluorescence (DIF) of the anagen and telogen hair (outer root sheath) and perilesional skin in a case of pemphigus vulgaris and in this case they found a correlation between them.
Daneshpazhooh M, et al in their study have found relatively high sensitivity of DIF on plucked hair in Pemphigus Vulgaris patients and that it is a suitable alternative to the more invasive techniques of skin or mucosal biopsy
Golan D,et al in their study have found the presence of auto antibodies to intercellular substance of squamous epithelium in the sera of 46 patients suffering from pemphigus vulgaris. This study supports the fact that pemphigus vulgaris is an autoimmune disease Kumar S, et al in their study observed DIF positive fluorescence of intercellular cement substance (ICS) of the epidermis in all 22 cases (100%) of their study. This study demonstrates the value of DIF for a definitive diagnosis of pemphigus
6.3 Objectives of study:
1. To compare the efficacy of direct immunofluorescence (DIF) of skin biopsy and plucked hair follicle in diagnosis of pemphigus.
2) To evaluate the significance of direct immunofluorescence (DIF) of outer root sheath of plucked hair follicles in diagnosis of pemphigus.
MATERIALS AND METHODS:
7.1 Source of data:
It is a prospective study from October 2013 to may 2015. Punch biopsy of skin & plucked hair follicles of pemphigus patients with clinically active lesions are received from department of Dermatology. They are processed in the department of Pathology, Victoria Hospital, Bangalore as per standard protocol.
7.2Methods of collection of data:
A) STUDY DESIGN: .Prospective study.
B) PERIOD OF STUDY: October 2013 to may 2015.
C) PLACE OF STUDY:
Department of Dermatology and Department of Pathology, Victoria Hospital, Bowring & Lady Curzon Hospitals, Bangalore.
D) SAMPLE SIZE :
Minimum of 35 samples (inclusive of both Skin punch biopsy & plucked hair follicles).
E) INCLUSION CRITERIA:
Punch biopsy of skin & plucked hair follicle samples in patients with pemphigus having clinically active lesions.
F) EXCLUSION CRITERIA:
Punch biopsy of skin & plucked hair follicle samples in patients with pemphigus having clinically inactive lesions.
Skin biopsy & plucked hair follicle specimens are taken from the lesions in patients of pemphigus with clinically active lesions. It is then subjected to Direct immunofluorescence(DIF). Fluorescin labelled monospecific antisera rose against IgA, IgG, IgM and complement component C3 are used in frozen sections from skin biopsy. Hair samples are subjected to DIF without sectioning by directly mounting it on the slide. The immunofluorescence pattern of both will be examined under IF microscope. The results will be recorded in the study proforma.
H) ASSESSMENT TOOLS: Direct immunofluorescence examination of the Skin biopsy & Hair follicles for the presence of antibody of the IgG, IgA, IgM class & the complement C3 component between epidermal cells.
Difference between continuous variables among groups will be assessed by studentt-test.
Categorical variables will be compared using Chi square test and Fischer’s exact test. P value of < 0.05 is considered significant.
7.3 Does the study require any investigation to be conducted on patients or animals? If so please describe briefly.
Yes. The study involves skin biopsy & plucked hair follicles in patients with pemphigus.
An informed consent is taken from the patients for all the necessary procedures & investigations.
Has ethical clearances been obtained from ethics committee of your institution in case of 7.3?
Yes. Ethical clearance will be obtained from the Institutional Ethical Committee. Bangalore medical college & research institute.
LIST OF REFERENCES:
1) Elder DE, Johnson B, Elenitsas R. Lever's Histopathology of the Skin. 9th Edition. Lippincott Williams & Wilkins. 2005; 244-291.
2) Alan D,Irwine MD,Harper's Textbook of Pediatric Dermatology, Volume 1, 2, Third Edition. Blackwell Publishing Ltd .2011;91.1-91.25
3). R.mark’s Rockberg,s common skin diseases 19th edition.;Chapman & hall.1993;87-88
4). Rao R,Dasari K,Shenoi S,Balachandran C et al, Demonstration of pemphigus-specific immunofluorescence pattern by direct immunofluorescence of plucked hair International journal of Dermatology.2009 Nov;48(111):187-9
5). Kumaresan,Reena Rai,andV Sandhyaet al Immunofluoresence of the Outer Root Sheath in Anagen and Telogen Hair: An Aid to Diagnosis in Pemphigus Int J Trichology.2009 Jul-Dec;1(2): 138–139
6). Daneshpazhooh M,Asgari M,Naraghi ZS,et al A study on plucked hair as a substrate for direct immunofluorescence in pemphigus vulgaris.J Eur Acad Dermatol Venereol.2009 Feb;23(2):129-319
7).Golan D,Gilhar A,Shmuel Z,Moshkowitz M. et al Autoantibodies to epithelial cells (intercellular substance) and their correlation with clinical activity of pemphigus vulgaris.Dermatologica. 1984;169(6):339-41
8). Kumar S, Thappa DM, Sehgal S. Immunofluorescence study of pemphigus. North India J Dermatol. Aug1995;22(8): 571-5.