Original Research. Oral mucocutaneous lesions - Histopathological and immunofluorescence study Rameshkumar A et al

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1 Received: 18 th Septemer 2014 Accepted: 20 th Decemer 2014 Conflicts of Interest: None Source of Support: Nil Originl Reserch Orl Mucocutneous Lesions A Comprtive Clinicopthologicl nd Immunofluorescence Study Annsmy Rmeshkumr 1, Alex Kumrnthr Vrghese 2, Thyln Dineshkumr 3, Shheen Ahmed 4, Jnki Venktrmni 5, G Sugirthrj 6 Contriutors: 1 Professor, Deprtment of Orl Pthology, SRM Dentl College, Chenni, Tmil Ndu, Indi; 2 Professor, Deprtment of Orl Pthology, Pushpgiri College of Dentl Sciences, Thiruvll, Kerl, Indi; 3 Reder, Deprtment of Orl Pthology, SRM Dentl College, Chenni, Tmil Ndu, Indi; 4 Professor & Hed (Retired), Deprtment of Orl Pthology, Tmil Ndu Government Dentl College nd Hospitls, Chenni, Tmil Ndu, Indi; 5 Professor & Hed (Retired), Deprtment of Dermtology, Mdrs Medicl College nd Hospitls, Chenni, Tmil Ndu, Indi; 6 Senior Lecturer, Deprtment of Orl Pthology, SRM Dentl College, Chenni, Tmil Ndu, Indi. Correspondence: Dr. Rmeshkumr A. SRM Dentl College, Bhrthi Sli, Rmpurm, Chenni , Tmil Ndu, Indi. Phone: Emil: How to cite the rticle: Rmeshkumr A, Vrghese AK, Dineshkumr T, Ahmed S, Venktrmni J, Sugirthrj G. Orl mucocutneous lesions A comprtive clinicopthologicl nd immunofluorescence study. J Int Orl Helth 2015;7(3): Astrct: Bckground: Orl is often ffected y mny diseses including mucocutneous disorders. The dignoses of these disorders re primrily sed on history, clinicl fetures, nd histopthology. For the pst few yers immunofluorescence techniques emerged s n importnt tool to study the pthogenesis nd in the dignosis of orl mucocutneous nd vesiculoullous disorders. The present study ws designed to crry out retrospective nd prospective nlysis of orl mucocutneous lesions to elucidte the clinicopthologic fetures nd its immunofluorescence findings. Mterils nd Methods: A totl of 70 sujects with orl mucocutneous lesions were retrieved from the orl pthology files of Tmil Ndu Govt. Dentl College nd their clinicl fetures were evluted, nd the histopthology ws lso evluted with the help of hemtoxylin nd eosin stined sections. For the prospective study, iopsy from 12 sujects with orl mucocutneous lesions ws sujected to routine histopthologicl exmintion nd DIF to evlute the consistency of the methods. Results: In the retrospective nlysis of 70 sujects with orl mucocutneous lesions in reltion to clinicl fetures nd histopthology, most of the findings were similr to the previous studies except for few criteri like mle predilection in lichen plnus nd mucous memrne pemphigoid (MMP) nd more prevlence of pemphigus vulgris thn MMP (2:1). In the prospective nlysis of 12 sujects, the histopthologicl dignosis ws consistent with DIF study in 66% of cses. Conclusion: The dignostic efficiency of orl mucocutneous lesions cn e improved y modern tools like DIF studies in ddition to trditionl methods like clinicl nd histopthology. Key Words: Histopthology, immunofluorescence, mucocutneous lesions, orl Introduction Orl is often ffected y mny diseses including mucocutneous disorders. The dignoses of these disorders re primrily sed on history, clinicl fetures, nd histopthology. The orl mucocutneous lesions cuse dignostic prolems, s these lesions resemle ech other, nd routine tissue iopsies my only offer dignosis of non-specific inflmmtion. 1 For the pst few yers, immunofluorescence techniques emerged s n importnt tool in detecting the pthogenesis nd dignosis of orl mucocutneous lesions (nd vesiculoullous disorders). Two types of immunofluorescence techniques tht commonly followed re direct immunofluorescence (DIF) nd indirect immunofluorescence (IIF). In some diseses, IIF studies hve reveled circulting utontiodies 2,3 wheres DIF studies hve reveled immunogloulins, complement components nd other protein sustnces within the ffected tissue. 4 These circulting nd tissue-ound sustnces my hve role in the immunopthogenesis of these diseses. In ddition detection of these immune rective sustnces y DIF ids in the dignosis of mny skin nd l diseses.¹ The presence of chrcteristic fluorescent ptterns in DIF cn definitely estlish the dignosis of pemphigus nd pemphigoid nd strongly indicte the likelihood of lichen plnus (LP) nd lupus erythemtosus. The sence of these fluorescent ptterns rules out these conditions, therey strengthening the dignosis of other orl l diseses. The results of DIF re sufficiently useful in the dignosis of chronic ulcertive diseses of the orl.¹ Bsed on these fcts, prospective study of orl mucocutneous lesions ws designed to evlute the consistency of the histopthology with DIF. A retrospective nlysis of the prevlence of orl mucocutneous lesions nmely orl LP (OLP), discoid lupus erythemtosus (DLE), pemphigus vulgris (PV), nd mucous memrne pemphigoid (MMP) were lso studied with regrds to demogrphic detils 59

2 nd histopthologicl fetures (hemtoxylin nd eosin stined sections) tht re chrcteristic of ech disese were evluted. Mterils nd Methods Twenty consecutive sujects in ech of OLP, DLE, PV, nd ten consecutive suject of MMP (totl 70 sujects) were retrieved from the orl pthology files of Tmil Ndu Govt. Dentl College nd reviewed for slient fetures including ge, gender, nd introrl site of the lesion. Hemtoxylin nd eosin (H nd E) stined sections of ll the 70 sujects were reviewed, nd the histopthologicl ptterns were evluted (retrospective study). Twelve sujects with chronic or recurrent ulcertive or erosive or vesiculoullous diseses of the orl were included for the prospective study. Biopsy specimens were tken from the perilesionl re in ll the 12 sujects. Specimens were exmined y routine histopthologicl methods (H nd E) nd DIF. Specimens for DIF study were quickly wshed in norml sline nd stored immeditely in Michel s medi nd trnsported to the lortory. Results Retrospective evlution The retrospective study comprised of 70 sujects - 20 sujects in ech of OLP, DLE, nd PV nd 10 sujects of MMP. The following significnt results were derived (Tle 1). OLP Eleven mles ffected ginst 9 femles (1:0:81). Fifth decde ws predominntly involved - 30% (6 sujects). Youngest involved ws 11 yers, nd oldest ws 72 yers, men ge yers. Buccl ws involved in 85% of the suject. Histopthologiclly, sl cell degenertion nd suepithelil nd of chronic inflmmtory cells were seen in 100% of sujects. PV Seven mles ffected ginst 13 femles, sixth decde ws predominntly involved - 45% (9 sujects). Youngest individul ws 33 yers, nd oldest ws 78 yers men ge - 52 yers. Buccl ws involved in 60% of sujects. Histopthologiclly, epithelium ulcerted in 40% of sujects, intct sl cells ttched to connective tissue seen in 100% of sujects, cntholytic (Tznck) cells seen in 85% of sujects nd dense chronic inflmmtory cells seen in the connective tissue in 55% sujects. MMP Seven mles ffected ginst 3 femles (1:0.42) fifth nd sixth decdes were predominntly involved - 60% (6 sujects). Youngest individul ws 33 yers, nd oldest ws 70 yers, men ge - 48 yers. Gingiv ws involved in 60% of sujects. Nikolsky s sign positive in 40% of sujects. Histopthologiclly, suepithelil split ws seen in 100% of the sujects, 40% showed inflmmtory cells nd red lood cells in the split, diffuse chronic inflmmtory cells seen in the connective tissue in 90% of sujects nd incresed vsculrity seen in 50% of sujects. DLE Five mles ffected ginst 15 femles (1:3), third nd fourth decdes were predominntly involved - 45% (9 sujects). Tle 1: Retrospective nlysis showing the demogrphic nd histopthologicl detils. Dignosis n Age (yers) Sex Site Epithelium Connective tissue Buccl Plte Lip Kertosis Less thn 20 yers Mle Femle Right Left Both Soft Hrd Upper Lower Tongue Gingiv Hyperkertosis Prkertiniztion Orthokertiniztion Hyperplstic Atrophic Ulcerted Intrepithelil split Suepithelil split Tzncks Bsl cell genertion Swtooth retepegs Chronic inflmmtory cells Melnin incontinence Perivsculr inflmmtory cells Cleft Lichen plnus Pemphigus vulgris MMP Discoid lupus erythemtosus MMP: Mucous memrne pemphigoid 60

3 Orl mucocutneous lesions - Histopthologicl nd immunofluorescence study Discussion Mucocutneous disorders (LP, DLE, PV, nd MMP) usully present themselves s chronic recurring ulcertions or vesiculoullous lesions, cusing ptients to seek dignosis nd tretment. Though histopthology remins the gold stndrd for the dignosis, in the pst few yer immunofluorescent techniques emerged s vlule tool in the dignosis of ulcertive nd vesiculoullous mucocutneous disorders.¹ Youngest individul ws 17 yers, nd oldest ws 70 yers men ge - 42 yers. Lip ws involved in 85% of sujects. Histopthologiclly, sl cell degenertion ws seen in 80% nd perivsculr chronic inflmmtory cells in 85% of sujects. Prospective evlution 12 cses of orl mucocutneous lesions were sujected to routine histopthologicl nd DIF nlysis. Histopthology showed 3 cses of LP, 4 cses of MMP nd 5 cses were dignosed s non-specific chronic inflmmtory lesions. In DIF study - 5 sujects were negtive. 3 were reported s OLP (Figure 1), 2 were interpreted s PV (Figure 2) nd 1 ech of MMP (Figure 3) nd LP pemphigoides (Figure 4). Histopthologicl dignosis consistent with DIF in 8 cses (66%) (3 OLP, 1 MMP nd 4 non-specific inflmmtion). LP is predominntly disese of the middle-ged nd elderly with ge rnging etween 30 nd 70 yers.5 In the present study lso, 65% of the ptients were in the sme ge groups. LP ffects oth the sexes, lthough occsionlly some survey hve suggested mle predominnce, mjority suggested femle predilection.6,7 The slight mle predilection in the present study (M: F=1:0.81) my e due to the smll smple size. Similr to the most of the studies in the literture,5 the present study lso showed uccl l predominnce. Similr to most of the studies,6 histopthology of the present study lso showed hyperorthokertosis, sl ll degenertion, nd suepithelil nd of chronic inflmmtory cells in 60%, 100%, nd 100% of the sujects, respectively. The histopthologicl dignosis nd DIF study were not consistent in four sujects in which one cse dignosed histopthologiclly s non-specific chronic inflmmtory lesion showed positivity for LP pemphigoides in DIF. Among the other three cses, one dignosed histopthologiclly s MMP ws negtive in DIF nd two cses which were dignosed s MMP histopthologiclly were found to show positivity for PV in DIF (Tle 2). Figure 3: (Mucous memrne pemphigoid) () Suepithelil cleft etween epithelium nd connective tissue (H nd E, 10), () direct immunofluorescence showing smooth, liner, nd continuous nd of C3 deposit long the sement memrne zone. Figure 1: (Lichen plnus), () Hyperkertosis, sl cell degenertion of epithelium long with suepithelil nd of chronic inflmmtory cells (H nd E, 10), () direct immunofluorescence showing firin deposition long the sement memrne zone. Figure 2: (Pemphigus vulgris) () Supr sl cleft with Tznck cells nd intct sl lyer (H nd E, 40), () direct immunofluorescence showing intercellulr spce deposition of immunogloulin G in the epithelium. Figure 4: (Lichen plnus pemphigoides) direct immunofluorescence showing liner deposit of C3 long the sement memrne zone. 61

4 Tle 2: Prospective nlysis showing the histopthologicl correltion with the immunofluorescence findings. Age (yers) Sex Skin lesions Site of orl lesions Clinicl dignosis Biopsy site Histopthologicl dignosis DIF findings DIF interprettion 52 M Gingiv MMP Gingiv MMP Strong ICS deposits of Pemphigus IgG in epithelium nd wek ICS deposits of C3 54 F Present Plte, lip, uccl Pemphigus Plte Non specific Negtive 59 M Plte Erosive LP Plte LP Strong discontinuous LP BMZ firous nd 27 F Buccl LP Buccl LP Strong continuous rgged LP BMZ nd of firinogen 47 M Buccl, gingiv LP/pemphigus Gingiv Non specific Negtive 47 M Buccl LP Buccl LP Modertely strong rgged LP BMZ nd of firinogen 32 M Present Buccl, tongue, floor, plte LP/pemphigus Buccl Non specific Wek discontinuous BMZ nd of IgG; wek BMZ Lichen plnus pemphigoides nd of C3; modertely strong BMZ nd of firinogen 48 M Buccl, tongue, Bullous LP/ Buccl MMP Wek BMZ nd of IgG MMP floor, gingiv pemphigus 45 M Buccl, tongue, Bullous LP/ Buccl MMP Negtive floor, gingiv pemphigus 52 F Gingiv MMP/ Gingiv MMP Wek ICS deposits of IgG Pemphigus desqumtive gingivitis 17 F Present Buccl, tongue, Erythem Tongue Non specific Non specific floor, lips multiforme 30 F Present Buccl, tongue, plte, lips, gingiv Bullous pemphigoid Plte Non specific Non specific DIF: Direct immunefluorescence, MMP: Mucous memrne pemphigoid, IgG: Immunogloulin G, BMZ: Bsement memrne zone, LP: Lichen plnus DLE is disese commonly seen in the third nd fourth decdes. It is lso considerly more common in women thn men. 6,8 The present study lso showed most of the sujects in the third nd fourth decdes (45%) with mle:femle rtio of 1:3. As with the other studies, 8,9 lip is the most common site of involvement in the present study (85%). A survey of literture showed tht DLE lesions re chrcterized y hyperkertosis, sl cell degenertion, perivsulr lymphocytic infiltrte, nd nd like inflmmtory cells in the lmin propri. 10 The present study lso showed sl cell degenertion, suepithelil nd of chronic inflmmtory cells, nd perivsculr chronic inflmmtion in 80%, 55%, nd 85%, of the sujects, respectively. Epithelil trophy lternting with hyperplsi showed high dignostic vlue for DLE ginst LP. 10 This feture ws seen in 35% sujects of the present study. PV predominntly seen etween third nd seventh decdes nd femles were commonly involved. In the present study lso 96% of the sujects were etween third nd seventh decdes nd femle to mle rtio ws 1:0.53. As with the mjority of the studies 6,8,11 in the present study lso uccl ws ffected in 60% of the sujects. The formtion of ull is entirely intrepithelil just ove the sl lyer, 8 Tznck cells nd intct sl cells ttched to connective tissue were seen in 80%, 85%, nd 100% of the sujects, respectively. These findings were in ccordnce with other studies. In contrst to other studies, 8,11 present study showed dense chronic inflmmtory cells infiltrte in 55% of sujects. MMP usully ffects older persons (fifth-eighth decde), femles re commonly involved (F:M: 2:1) 6 In the present study lso 70% of the ptients were in the ge group of fifth to eighth decdes. More numer of mles (70%) ffected in the present study my e ttriuted to the smll smple size. Gingiv is the most common site of involvement followed y uccl. 8,12 In the present study lso gingiv ws involved in 60% of sujects nd uccl ws lso eqully involved. The su epithelil split nd diffuse chronic inflmmtory cell infiltrte were seen in 100% nd 90% of the sujects, this is in ccordnce with the previous studies. 8,12 In our prospective study of 12 sujects of orl l lesions y DIF, 5 were negtive, 3 were LP, 2 were PV, nd one ech of MMP nd LP pemphigoides. Histopthologicl dignosis ws consistent with DIF in 66% of sujects, nd this ws in ccordnce with the previous studies y Dniels nd Crmen.¹ Histopthologicl nd DIF results were similr in 3 cses of OLP, nd one cse of MMP nd 4 sujects which were found to e nonspecific inflmmtion were lso found to e negtive in DIF study. Negtive results in DIF will help us to rule out those diseses which produces specific fluorescent ptterns nd therey strengthening the dignosis of other orl l lesions. 62

5 Though in some cses histopthology is not consistent with DIF oth were le to offer dignosis in 2/3 of the sujects. Erly dignosis of MMP is importnt since the prevlence of concurrent eye lesions is greter with orl lesions. DIF is more specific nd sensitive thn histopthology nd electron microscopy in dignosing MMP Immunofluorescence nlysis is lso helpful tool in monitoring therpeutic responses of these lesions y nlyzing the lesion specific utontiodies. 16 Conclusion Although the histopthology remins the gold stndrd for the dignosis of most of the diseses, it is inferred from this study tht not ll lesions re menle to definitive dignosis. Due to the ccurcy of DLF study in dignosing diseses, its report is tken s the finl dignosis, which scertins the course nd prognosis of the disese. However, the consistency of the DLF in this study cnnot e sustntited due to the limited smple size which is ttriuted to cost effectiveness of the DLF technique. Acknowledgments The primry uthor would like to cknowledge the vlule inputs y KA Annsmy nd A Vthsl nd lso would like to thnk A Pnneerselvm, A Alguselvm, P Kunthvi, A Girij, Dr. S Shnthy, P Chndrnu nd P Sthy in prepring the mnuscript nd proofreding. References 1. Dniels TE, Qudr-White C. Direct immunofluorescence in orl l disese: A dignostic nlysis of 130 cses. Orl Surg Orl Med Orl Pthol 1981;51(1): Beutner EH, Chrozelski TP, Kumr V. Immunopthology of Skin, 3 rd ed. New York: John Wiley nd Sons; Mutsim DF, Adms BB. Immunofluorescence in dermtology. J Am Acd Dermtol 2001;45(6): Dyn S, Simmons RK, Ahmed AR. Contemporry issues in the dignosis of orl pemphigoid: A selective review of the literture. Orl Surg Orl Med Orl Pthol Orl Rdiol Endod 1999;88(4): Mlloglu N. Orl lichenplnus - A review. Br J Orl Mxillofc Surg 2000;38(4): Allen C, Neville B, Dmm D, Bouquot J. Orl nd Mxillofcil Pthology, 1 st ed. Phildelphi: W. B. Sunders Compny; Scully C, el-kom M. Lichen plnus: Review nd updte on pthogenesis. J Orl Pthol 1985;14(6): Shfer WG, Hine MK, Levy BM. Text Book of Orl Pthology, 4 th ed. Phildelphi: W. B. Sunders Compny; Regezi JA, Sciu JJ. Orl Pthology, 2 nd ed. Phildelphi: W. B. Sunders Compny; Schiødt M. Orl discoid lupus erythemtosus. III. A histopthologic study of sixty-six ptients. Orl Surg Orl Med Orl Pthol 1984;57: Weinerg MA, Insler MS, Cmpen RB. Mucocutneous fetures of utoimmune listering diseses. Orl Surg Orl Med Orl Pthol Orl Rdiol Endod 1997;84(5): Fleming TE, Kormn NJ. Cictricil pemphigoid. J Am Acd Dermtol 2000;43(4):571-91; quiz Eschle-Meniconi ME, Ahmd SR, Foster CS. Mucous memrne pemphigoid: An updte. Curr Opin Ophthlmol 2005;16(5): Dniel E, Thorne JE. Recent dvnces in mucous memrne pemphigoid. Curr Opin Ophthlmol 2008;19(4): Chn LS. Oculr nd orl mucous memrne pemphigoid (cictricil pemphigoid). Clin Dermtol 2012;30(1): Knudson RM, Klji AN, Bruce AJ. The mngement of mucous memrne pemphigoid nd pemphigus. Dermtol Ther 2010;23(3):

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