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Indian Journal of Human Genetics
Medknow Publications on behalf of Indian Society of Human Genetics
ISSN: 0971-6866 EISSN: 1998-362x
Vol. 11, Num. 1, 2005, pp. 24-26

Indian Journal of Human Genetics, Vol. 11, No. 1, January-June, 2005, pp. 24-26

Original Article

Polymorphism of alpha-1-antitrypsin and association of S and Z alleles with duodenal ulcers

Sulekha S., Mathur B.P., Pratibha N.

Department of Genetics, Osmania University, Hyderabad

Correspondence Address: 33/426 Arjun Vihar, Defence Officers Enclave, New Delhi - 110010, shashilekh@indiatimes.com

Code Number: hg05002

ABSTRACT

BACKGROUND: Duodenal ulcers are mucosal erosions that penetrate into the muscularis propria of the duodenum. They are a result of an imbalance between aggressive and defensive factors. Various environmental factors like Helicobacter pylori infection, addictions to smoking and alcohol etc. and genetic factors have been reported to be associated with duodenal ulceration. Alpha-1-antitrypsin was studied for its role as a genetic marker and specific allelic association to protein functioning and alteration. Serum samples from 185 normal subjects and 210 duodenal ulcer cases were typed for the phenotypes following PAGE (polyacrylamide gel electrophoresis) and immunofixation using specific commercial antisera with appropriate staining protocols. In general, 'M' allele of alpha-1-antitrypsin was found to be predominant in healthy normal subjects, with the gene frequencies being 0.679 (M), 0.299 (Z) and 0.0214 (S). Whereas in duodenal ulcer cases, Z and S alleles were found to be predominant with a significant association of MS, ZZ and MZ phenotypes (χ2sub : 49.98) and the gene frequencies being 0. 113 (M), 0. 347 (Z) and 0.506 (S). Predominance of Z and S alleles indicates that these alleles may encode for reduced synthesis of alpha-1-antitrypsin, hence decreased neutralization of proteases like trypsin and chymotrypsin inhibited by alpha-1-antitrypsin, thereby resulting in ulcers. The study highlights the association of Z and S alleles of the potent protease inhibitor alpha-1-antitrypsin and also suggests its role as a genetic marker in ulcerogenesis.

Keywords: Protease inhibitor, tissue degradation, glycoprotein, genetic marker

INTRODUCTION

Duodenal ulcers are lesions on the mucosa that extend into the muscularis propria of the duodenum. They occur in the first part of the duodenum. An imbalance between aggressive (acid, pepsin) and defensive (mucosal defenses) factors is the primary event for the disruption of mucosal integrity leading to ulcerogenesis.

Duodenal ulcers have been reported to be a net result of environmental and genetic factors with the mode of inheritance being multifactorial.[1] Earlier studies reported an association of blood group O,[2] ABH non - secretor status,[3] HLA-B5 phenotype,[4] Gc 1-1 phenotype[5] with duodenal ulceration. Hyperpepsinogenemia (PG) was reported to be a sub-clinical risk factor in duodenal ulceration.[5],[6],[7] The role of Cathepsin E and alpha-1-antitrypsin as defensive factors in duodenal ulceration[5],[8] was also reported.

Alpha-1-antitrypsin, a major protease inhibitor that protects the mucosal tissue from degradation is found to be localized on chromosome 14 q 31. Trypsin and Chymotrypsin are the two proteases confirmed by earlier studies as etiological factors in duodenal ulceration that are known to be inhibited by alpha-1-antitrypsin. Recent studies have shown the association of S and Z alleles of alpha-1-antitrypsin with deficiency of the enzyme.[9] Thus, the study envisages into the polymorphic variation of alpha-1-antitrypsin and its role as a genetic marker in ulcerogenesis.

MATERIALS AND METHODS

With the approval of the local ethics committee, the study envisaged the analysis of blood samples from 210 endoscopically confirmed duodenal ulcer patients, and 185 normal subjects (without prior medication) that were collected from the Gastroentererology unit of Osmania General Hospital, Hyderabad, by visiting the hospital twice a week (Mondays and Thursdays) during the period of study (1998-2001). No patient was excluded from the study. Apart from blood samples, epidemiological data regarding the socio economic stata, familial history and food habits including addiction to smoking and alcohol were collected from each of the individuals of both the disease and control groups. The blood samples were allowed to coagulate and immediately centrifuged at 2000rpm for 5 minutes. The serum samples obtained after centrifugation were analysed for the phenotypes of alpha-1-antitrypsin, carried out on 7% polyacrylamide gels following polyacrylamide gel electrophoresis (PAGE).[10] The glass plates with polymerized separating gel were fixed to a vertical slab gel electrophoretic unit. The upper and lower tanks containing the tank buffer were connected to the anodic and cathodic ends of the DC power supply. 40 ml of serum with 20ml of bromophenol blue indicator were loaded in the sample wells. Electrophoresis was conducted at 20 mA constant current at 4oC for 4-5 hours. The gel was then subjected to immunoblot with alpha-1-antitrypsin antisera (Sigma grade) and was incubated overnight at 4oC that was later washed and stained[11] with simultaneous destaining until distinct bands of alpha-1-antitrypsin were visible on a clear background.

The data was computed and analysed for gene frequencies and Woolf′s test of association was carried out to identify any specific electromorphic association with duodenal ulcers.

RESULTS

The frequency distribution of alpha-1-antitrypsin electromorphs in control and disease groups presented in [Table - 1] indicates a general predominance of M allele in normal subjects and Z and S alleles in disease condition. In normal subjects, 44.0% were of MM type, followed by MS (16.0%), MZ (12.0%) and ZZ (8.6%) phenotypes. While in duodenal ulcer cases, 32.0% were of MS, 23.3% were ZZ and 22.0% were of MZ phenotypes.

Test of association of alpha-1-antitrypsin phenotypes in duodenal ulcer cases is presented in [Table - 2]. Relative risks estimates (RR) of various phenotypes were calculated of which MM phenotype when compared with MZ, MS and FM revealed significance with the relative risks being 0.09 (χ2: 49.98*) and ZZ phenotype when compared with SZ and MZ phenotypes had relative risks of 1.2 (χ2: 0.28) with no significant association. Similarly, homozygotes when compared with heterozygotes revealed a significant predisposition of heterozygotes to duodenal ulcer, with the relative risk being 0.46 (χ2: 13.84*) (*: P <0.01).

The allelic frequencies of M, S and Z phenotypes of alpha-1-antitrypsin are presented in [Table - 3]. The gene frequencies were calculated and were observed to be 0.679(M), 0.299(Z) and 0.0214(S) in control subjects and 0.113(M), 0.347(Z) and 0.506(S) in duodenal ulcer cases.

DISCUSSION

The predominance of S and Z alleles in the disease condition compared to the control group, observed in the present study suggests that these alleles may encode for reduced synthesis of alpha-1-antitrypsin, which can be strengthened by earlier studies.[9] It can also be thought on the lines that, any abnormalities of the liver and its metabolism may also lead to reduced synthesis of this glycoprotein since the liver is the site of alpha-1-antitrypsin synthesis. As glycoproteins work to protect the surface of the gastric mucosa against the effects of acid and toxic metabolites, the alteration may thereby result in failure to render mucosal cytoprotection. These conditions may make the gastric mucosa more susceptible to autodigestion by tissue degrading proteases like trypsin and chymotrypsin due to the inability of alpha-1-antitrypsin to form molar complexes with them. Hence, decreased protection conferred to the gastric mucosa from the proteolytic enzymes highlights the role of alpha-1-antitrypsin as a genetic marker in ulcerogenesis. This also indicates an imbalance between aggressive and defensive mechanisms. Thus, predominance of S and Z alleles of alpha-1-antitrypsin may help as a pre-clinical marker to aid in early treatment. Although, the study implicates to explore the role of other genetic and environmental factors influencing the process of ulcerogenesis.

ACKNOWLEDGEMENT 

We acknowledge Indian Council of Medical Research (ICMR) for providing financial assistance to accomplish the present study.

REFERENCES

1.Kurata JH, Haile BM, Elashoff JD. Sex difference in peptic ulcer disease. Gastroenterol 1985;88:96-8.  Back to cited text no. 1    
2.Mourant AE, Koper AC, Sobekaz KD. Blood groups and Diseases: A study of association of diseases with blood groups and other polymorphisms. Oxford: University press; 1978.  Back to cited text no. 2    
3.Clarke CA. Heredity and the gastrointestinal tract. In : Genetics for the clinician. Philadelphia: Blackwell Scientific Publication; 1964;Ch.9:113.  Back to cited text no. 3    
4.Ellis A, Woodrow JC. HLA and duodenal ulcer. Gut 1979;20:760-2.  Back to cited text no. 4    
5.Venkateshwari A. Duodenal Ulcers: A study on genetic and biochemical aspects. PhD Thesis. Hyderabad, India: Osmania University; 1998.  Back to cited text no. 5    
6.Rotter JI, Peterson G, Samloff IM, Mc Conell RB, Ellis A, Spons MA, et al. Genetic heterogeneity of hyperpepsinogenemic I and normopepsinogenemic I duodenal ulcer. Ann Intern Med 1979;91:372-7.  Back to cited text no. 6    
7.Taylor IL, Byrne WJ, Christie D, Ament ME, Walsh JH. Effect of individual L-amino acids on gastric acid secretion and serum gastrin and pancreatic polypeptide release in humans. Gastroenterol 1982;83:273.  Back to cited text no. 7    
8.Rajshree K. The role of proteolytic and pancreatic enzymes and serum proteins in the etiopathogenesis of Duodenal Ulcers. PhD Thesis. Hyderabad, India: Osmania University; 1994.  Back to cited text no. 8    
9.Shahid A, Siddiqui AA, Sultana T, Qureshi H, Waqar MA, Zuberi SJ. Determination of alpa-1-antitrypsin genetic deficiency in duodenal ulcer by polymerase chain reaction. J Pak Med Assoc 2002;52:545-8.  Back to cited text no. 9    
10.Davies BJ. Disc electrophoresis II. Method and application to human serum proteins. Ann N Y Acad Sci 1964;21:404-32.  Back to cited text no. 10    
11.Wilson CM. Commassie Staining. Methods in Enzymology . 1983;91:236-47.  Back to cited text no. 11    

Copyright 2005 - Indian Journal of Human Genetics

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