A, Zuberi Mohammad

J, Sarwar

Shahid, Anjum

Siddiqui, Anwar Ali

Sultana, Taranum

Waqar,Huma Qureshi, Waqar,Huma Qureshi Waqar,Huma Qureshi

English

eCommons@AKU

eCommons@AKUDepartment of Biological & Biomedical Sciences Medical College, PakistanSeptember 2000Low serum alpha 1 antitrypsin in duodenal ulcer--afamily studyAnjum ShahidMedical Centre, KarachiSarwar JMedical Centre, KarachiZuberi Mohammad AMedical Centre, KarachiWaqar,Huma Qureshi Waqar,Huma Qureshi Waqar,Huma QureshiMedical Centre, KarachiAnwar Ali SiddiquiAga Khan University, anwar.siddiqui@aku.eduSee next page for additional authorsFollow this and additional works at: https://ecommons.aku.edu/pakistan_fhs_mc_bbsPart of the Gastroenterology Commons, Genetic Processes Commons, Medical EducationCommons, and the Medical Genetics CommonsRecommended CitationShahid, A., J, S., A, Z. M., Waqar,Huma Qureshi, W. W., Siddiqui, A. A., Sultana, T. (2000). Low serum alpha 1 antitrypsin in duodenalulcer--a family study. JPMA. The Journal of the Pakistan Medical Association, 50(9), 318-320.Available at: https://ecommons.aku.edu/pakistan_fhs_mc_bbs/254AuthorsAnjum Shahid; Sarwar J; Zuberi Mohammad A; Waqar,Huma Qureshi Waqar,Huma Qureshi Waqar,HumaQureshi; Anwar Ali Siddiqui; and Taranum SultanaThis report is available at eCommons@AKU: https://ecommons.aku.edu/pakistan_fhs_mc_bbs/254Low Serum Alpha 1 Antitrypsin in Duodenal Ulcer - A FamilyStudyAnjum Shahid,Sarwar J. Zuberi,Mohammad A. Waqar,Huma Qureshi ( PMRC Research Centre, Jinnah PostgraduatcMedical Centre, Karachi. ) Anwar A. Siddiqui,Taranum Sultana  ( Department of Biochemistry, The Aga Khan University, Karachi. ) Introduction Genetic association of a disease is often recognized due to its occurrence in families. Duodenal ulcerlike many common diseases has a variable age of onset, expression and familial aggregation   1. Threelines of evidence support a genetic role: family studies, twin studies and blood group studies. Familyaggregation is commonly associated with early onset (<30 years) and blood group 0 with late onset ofsymptoms2. The disease has a genetic component that predisposes and an environmental factor thathelps in its manifestation1.Alpha-l antitrypsin (alpha I AT) deficiency is an inherited metabolic disorder associated with not only asevere reduction of alpha- AT in blood but also aminoacid substitution of alpha- I AT due to genevariations3. Subjects With alpha I AT, deficiency4  particularly its Z allele are moc prone to developduodena ulcers5. This association may be a contributory factor towards the aggravation of the diseaseand this perception becomes even stronger when other risk factors for duodenal ulcer like helicobacterpylon are absent among the family members. Z mutation is the most common genetic defect in alphaAT deficiency and its screening is commonly done by Isoelectric focusing (IEF). However, for accurateidentification genetic screening has been recommended which includes detection of inherited traits bymeasurements of enz\\me activities in blood, the presence of a specific gene or a specific mutation4.We, therefore have applied both the methods and have found that polymerase chain reaction (PCR)based method is an effective way to confirm the phenotypes6.Case Report In Pakistan, duodenal ulcer is infrequent in females especially those who are not taking non-steroidalanti-inflammatory drugs (NSAID)7. The proband. a female developed ulcer symptoms at the age of 50years. Upper gastrointestinal (GI) endoscopy at that time showed streaks of esophagitis with noabnormality in stomach and duodenum. After a year she was re-endoscoped as her symptoms hadworsened. This time apart from esophagitis, a large anterior wall duodenal ulcer with contact bleedingwas seen, she was treated with Ran itidine (300 mg) at night for a month. Repeat endoscopy after 4weeks showed a healing duodenal ulcer (smaller than before). The drug was continued for 8 weeks andthen reduced to half the dose at bed time for another 8 weeks. Patient was asymptomatic for almost 9months, when her symptoms recurred along with epigastric tenderness. Ranitidine was given foranother 8 weeks, after which she again became asymptomatic. She had multiple relapses since then,each treated with H2 receptor antagonists. On last relapse she also complained of vomiting andinability to retain food. Upper GI endoscopy showed linear streaks of severe esophagitis with bilereflux, elongated stomach with billious gastric residue and pyloric stenosis. At this stage patient refusedsurgery and started self-medication. Tests for helicobacter pylon were negative. It was thereforeconsidered appropriate to screen for various genetic markers i.e.. alpha 1 AT, serum pepsinogen andblood group. Written consent was obtained from all the patient and her family members and anapproval was granted by the ethical committee of the Jinnah postgraduate medical centre, Karachi tostudy genetic markers. Serum sample drawn from patient was subjected to radial immunodiffusion(RID)6 and (IEF)8 for the estimation of serum alpha-I AT levels and determination of phenotypesrespectively. The concentration of alpha-I AT by RID was less than 50% of the normal level (any valuehas than 2 gm/I was considered below normal). Phenotyping revealed a deficiency variant that wasinferred to be ZZ phenotype as no prominent protein bands were visible in the area designated foralpha-I AT indicating the presence of a partial to total alpha-I AT deficiency. Further confirmation ofphenotyping was done by deoxyribonucleic acid (DNA) analysis techniques. DNA was extracted9 anda combination of PCR and restriction enzyme digestion10 was then applied to confirm the identity ofdeficient variant. Her blood group was AB and serum pepsinogen concentration using the method ofEdwards et al11 was above the normal expected range i.e., 179 units/mI (normal range: 60-160units/mI).The family history of the proband showed that her parents were first cousins, among the immediatefamily, one of her brother had a duodenal ulcer. All the other siblings and offsprings screened for serumalpha I AT by the same methods were healthy adults with no signs and symptoms of duodenal ulcer.None of the sibling and offsprings had helicobacter pylor as judged by the CLO test12.Serum samples from three siblings, two daughters and one son were analyzed. The results are presentedin the accompanying Table and Figure 1.One of the sibling had a low level on the partigen plate and IEF showed a pattern that looked like aheterozygous variant that was not clearly resolved into MM or MZ. Other two siblings showed normalvariants of M by IEF.One of the daughters also had a normal variant and normal concentration of alpha-I-AT whereas the sonand the other daughter and low levels of alpha I AT. This daughter also exhibited a pattern on IEF thatappeared indistinct. The son demonstrated a pattern on LEF which was very similar to that of theproband and clearly exhibited a pattern of ZZ type. To confirm the IEF results, particularly in one ofthe siblings and daughter with low alpha 1 AT, molecular techniques were applied which not onlyprovides early identification but also distinguishes heterozygous and homozygous alpha-i antitrypsindeficient13. The results of the typing for the Z mutation more clearly established the presence of Zmutation in the proband and her son. The relatively unresolved phenotyping carried out by IEF of onesibling and a daughter who had low alpha-I AT levels with indistinct patterns showed that the daughterand the sibling both had MM phenotypes. as judged by PCR and subsequent restriction digestion. Thusnormal persons and those homozygous for the Z mutation were distinguished unambiguously byapplying PCR based method. All the subjects with normal MM phenotype believed to have the normalsequence as their amplified product by PCR and subsequent digestion with restriction enzyme Taq Iproduced a DNA fragment of 64 base pair, that is only observed in the absence of a mutation. While theproband and her son’s DNA treated in the same way manifested Z mutation by showing a final productat the 86 base pair position as analyzed on agarose gel (Figure 2).In our experience IEF should be used as the screening method and the DNA based techniques as theverification of cases where serum concentration is low and IEF does not clearly discriminate thePhenotypes.Discussion Duodenal ulcer is a common gastrointestinal problem in this part of the world and a search for thecausative and aggravating factors is in progress. Relatively few evidences support a positive correlationbetween alpha1 AT deficiency and duodenal ulcer and these are mostly of preliminary nature4,5. Wewere able to identify an ulcer patient, with low serum alpha-I AT with a deficient variant on LEF, whichwas further confirmed by molecular techniques. This patient also had a raised serum pepsinogen. Theseobservations suggested the etiology to be of genetic origin. Such an association with genetic markersdemonstrates the importance of genetic factors and also serves in the detection of geneticallypredisposed persons1. Recent advances in molecular genetics have improved our understanding of thedisease and these scientific advances have important implications in the diagnosis, treatment andscreening of patients with genetic diseases and their families14. Considering the patient’s follow upwhich was almost over six years and in-spite of treatment with H2 receptor antagonists, she hadrelapses of ulcer. The genetic origins of ulcer disease could have been one of the factors responsible fora poor response to therapy leading to frequent relapses in the absence of H. pylon. It is possible thatdue to alpha- I AT deficiency, activity of proteolytic enzymes remained unchecked and resulted infrequent recurrences15. In the present study more than 50% of the offsprings and one sibling alsomanifested low levels of alpha-I AT. All these were healthy adults with no signs and symptoms ofduodenal ulcer. This situation is very much similar to an earlier reported study16 where by using serumpepsinogen as a genetic marker in duodenal ulcer; it was suggested that the family members of patientswith raised serum pepsinogen should be screened to identify those carrying the trait, so that they couldbe protected from such factors that may induce and/or aggravates the disease process. The samehypothesis may hold true in the present study where it could be suggested that family members ofduodenal ulcer patient carrying that trait are at an increased risk for duodenal ulcer.References 1. Hahibullah CM, Radha V The genetics of peptic ulcer disease, Trop. Gastroeni. 1985,6.1 32-41.2. Lam SK. Epidemiology and genetics of peptic ulcer Gastroenterol Jpn., 1993.28 45-57.3. Miv ak K. Wakashima M Liver cirrhosis associated with alphaII antitrypsin deficiency. NipponRinsho . 994:52:21 5—21.4. Kishore N Alpha-I AT deficiency in duodenal ulcer. Trop. Gastroent., 1980. 193-96.5. Shahid A, Siddiqu AA. Luberi SJ, et al, Duodcnal ulcer and alpha-I antitrypsin deficiency. J.Gastroent. I Hepatol.. 1 1993:8,505-507.6. Mancini G, Carbonarci AO, Herematis JR Immunochemical quantiation of’ antigens by single radialiminunodiffusion Immuno. Chem,, 1965:2:235-54.7. Ahmed W, Qureshi It, Alani SF. et at, Patient of \'duodenal ulcer in Karachi. J. Pak. Med. Assoc..1990;40:212-15.8. Jcppsson JO. Franzen 13 Typing of genetic variants of alpha-I AT by electrofocusing Clin Client..1982;28:219-25.9. Miller SA, Dykes DD, Poleskv H IF. A simple salting out procedure for extracting human l)NA fromhuman nucleated cells. Nucl Acid Res., 1988: 16: 12 15-18.10. Andresen BS, Knudsen I. Jensen PKA, et at. Two novel non radioactive polymerase chain reactionbased assays of dried blood spots, genoinic DNA or whole cells for fast reliable detection of Z and Smutations in the alpha-l AT gene. Clin Chem 1992;38:2100-2107.11. Edward K, Jepson RP, Wood KF. Modified serum pepsinogen. In Harold Varlcv, cd. PracticalClinical Biochemistry 5th edition. London, William Heinemann 1980, pp. 1003—1004.12. Cliodna MM. Mc-Nutty JC, Den Off JS. et al. Detection of Campylobacter pylor by biopsy uneasetest An assessment in 1445 patients. Gut. 1989;30:1058-62.13. Lain CWK, Pang CP, I’oon PMK. et at, Rapid screening for alpha-I antitrypsin Z and S mutations.Clin. Chen,. 1997;43,403-404.14. Tung BY. Kowdley K \\‘. Inherited liver diseases affecting the adult. Gastroent.. 1996;4:245-61.15.  Novis BH, Bank S. Young GO, et at. Chronic pancreatitis and alpha—I-AT. Lancet, I 965;2:748-49.16 .Habibullah CM, Alt MM. lsliaq M. et al. Study of duodenal ulcer disease in 100 families using totalserum pepsinogen as a genetic marker. Gut,1984;25:1380-83.

Low serum alpha 1 antitrypsin in duodenal ulcer--a family study

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Low serum alpha 1 antitrypsin in duodenal ulcer--a family study

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