Simplified microsatellite instability detection protocol provides equivalent sensitivity to robust detection strategies in Lynch syndrome patients

Objective: Germline mutations in mismatch repair (MMR) genes cause Lynch syndrome (LS). LS is an inherited disease, and an important consequence of MMR deficiency is microsatellite instability (MSI) phenotype. MSI phenotype influences the efficacy of 5 fluorouracil (5-FU) chemotherapy. Reproducible, cost effective, and easy to perform laboratory tests are required to include MSI detection in routine laboratory practice. Evaluation of CAT25 as monomorphic short tandem repeat sequence enables CAT25 to be an efficient screening tool among hereditary nonpolyposis colorectal cancer (HNPCC) patients compared with other methods used currently. Methods: Based on Amsterdam II criteria, 31 patients in 31 families were shortlisted from a total number of 1,659 colorectal cancer patients. MSI status was examined in these patients using CAT25 and a commercially available Promega MSI five-marker-based detection system as well as immunohistochemical (IHC) staining of four important MMR proteins. Patients were scored as high microsatellite instable (MSI-H), low (MSI-L), or stable (MSS). MSI status determined by CAT25 single mononucleotide marker was compared with that of five mononucleotide markers, Promega commercial kit, and IHC method. Results: MMR protein deficiency was observed on 7/31 probands using IHC methodology and 6/31 categorized as MSI-H using commercial kit or CAT25 single marker. The sensitivity and specificity of the CAT25 single marker were the same as those detected by five-marker Promega commercial kit in our patients. Conclusions: Based on our results, the performance of the CAT25 single mononucleotide marker for MSI status determination in our HNPCC patients is the same as that of the five-marker-based commercial kit

Genetic linkage analysis of DFNB59 loci involved in autosomal recessive non-syndromic hearing loss (ARNSHL) in western provinces of Iran

Bilateral sensorineural hearing loss is one of the most frequent congenital defects, and it has the prevalence of one in thousand among the infants. The most cases of hearing impairment are nonsyndromic and about 80 percent of patients with sensorineural HE show authosomal recessive pattern. ARSNL is a very heterogenic defect for over 70 loci and 50 genes have been found. In this study, in order to prove the prevalence of DFNB59 mutations, we examined DFNB59 mutations in 54 families from 3 provinces of Iran including Hamedan, Kohgiluye& Boveir Ahmad and Chaharmahal Bakhtiari. The gene (PJVK) in this locus encodes Pejvakin protein (PJVK) and causes nerve stimulation to conduct. Selected families in this study have consanguinity and at least have two patients with the defect they also are negative in having the GJB2 mutations. Six markers were selected for this locus and linkage analysis was performed. However, after examining different families, it was revealed that none of them had linkage to the DFNB59 locus. Therefore, this study suggests that that Pejvakin mutations might not play an important role in causing hearing loss in these provinces

MT1XT20 single quasi-monomorphic mononucleotide marker for detection of microsatellite instability in iranian patients with hereditary nonpolyposis colorectal cancer (HNPCC)

Background: Colorectal malignancies with high microsatellite instability (MSI-H), either hereditary or sporadic, demonstrate better prognosis, altered response to fluorouracil (5FU) chemotherapy and altered operative approach. It is now recommended to perform MSI testing for all new cases of colorectal cancers regardless of being categorized as hereditary or sporadic. This study aimed to evaluate MT1XT20 mononucleotide marker in Iranian patients with hereditary nonpolyposis colorectal cancer (HNPCC). The samples were further characterized using Promega five-marker MSI testing panel and immunohistochemical (IHC) technique. Methods: MT1XT20 mononucleotide marker and commercially available kit (Promega, USA) incorporating five quasi-monomorphic markers were studied in 20 cases of HNPCC using polymerase chain reaction (PCR) technique. IHC was performed to evaluate the status of all four important mismatch repair (MMR) proteins, too. Findings: Eight (40%), seven (35%) and five (25%) cases showed MSI using Promega kit, IHC and MT1XT20, respectively. Among the markers included in Promega kit, BAT26 marker with instability in all 8 samples (100%) was the most instable marker. NR24 and NR21 markers showed instability in 7 cases (87.5%); BAT25 and MONO 27 markers were instable in 6 (75.0%) and 5 (62.5%) specimens, respectively. Conclusion: Although MT1XT20 is considered as a valid single marker in Italian population, it seems this is not hold true about the Iranian patients. Instead, BAT26 among the markers included in Promega MSI testing was shown instability in all 8 samples of MSI-H colorectal cancer (CRC). Therefore, it may be concluded that BAT26 alone is as efficient as the cohort of five markers in Iranian patients. © 2016, Isfahan University of Medical Sciences(IUMS). All rights reserved

Genetic linkage analysis of DFNB40 and DFNB48 loci in families with autosomal recessive non-syndromic hearing loss (ARNSHL) from western provinces of Iran

Background: Sensorineural hearing loss (SNHL) is the most common sensory disorder and 1 in every 500-1000 newborns is affected. Non-syndromic SNHL accounts for 70% of hereditary hearing loss and 80% of SNHL cases have an autosomal recessive mode of inheritance (ARNSHL). The Purpose of the recent study is genetic linkage analysis to determine the prevalence of DFNB40 and DFNB48 loci in studying families with ARNSHL from the western provinces of Iran. Methods: In this study, 60 families from 3 provinces of Iran involving Hamedan, Kohgiluyeh and Boyer-Ahmad and Chaharmahal and Bakhtiari with autosomal recessive non syndromic hearing loss were examined. The selected families in this study were consanguineous and had at least two patients. They also were negative for GJB2 mutations. Linkage analysis was performed by using 6 markers short tandem repeat (STR) for the DFNB40 locus and 7 markers STR for the DFNB48 locus. Findings: After examining different families, it was revealed that none of them showed linkage to the DFNB40 and DFNB48 loci. Conclusion: The recent study suggests that DFNB40 and DFNB48 loci might not play an important role in causing hearing loss in the mentioned provinces. However, further studies are necessary to determine more precisely the role of these loci in the Iranian population. © 2016, Isfahan University of Medical Sciences(IUMS). All rights reserved

Genetic linkage analysis of the DFNB63 locus in families with autosomal recessive nonsyndromic hearing loss from Hamadan and Kohgiluyeh and Boyer-Ahmad Provinces, Iran

Background: Hearing loss is a sensorineural impairment and is one of the most widespread congenital impairments with a prevalence of one in thousand among children. Studies have shown that 50 percent of congenital hearing loss have genetic causes and the remaining 50 percent are due to environment and unknown reasons; in addition, it is noted that this impairment is very heterogeneous. Almost 70 percent of cases are nonsyndromic with hearing loss presenting as the only impairment. About 80 percent of this type of hearing loss is inherited in recessive manner (ARNSHL). In this study, we determined the role of DFNB63 locus in a series of families in two western provinces of Iran. Methods: In this descriptive-laboratory study, to determine the prevalence of DFNB63 mutations in western provinces of Iran, we studied 150 individuals from 30 families in Hamadan and Kohgiluyeh and Boyer-Ahmad provinces. The selected families in this study were consanguineous, had at least 2 patients, and were negative for GJB2 mutations. Linkage analysis was performed using six appropriate short tandem repeats (STR) markers. Findings: With linkage analysis of selected families, no family was shown to be linked to the DFNB63 locus. It was shown that the LRTOMT mutations played no role in causing hearing loss in the studied families. Conclusion: The present study suggests that LRTOMT mutations may not be clinically important in causing autosomal recessive nonsyndromic hearing loss in the investigated provinces. © 2015, Isfahan University of Medical Sciences(IUMS). All rights reserved

Genetic linkage analysis of DFNB7/11 locus in patients with autosomal recessive non syndromic hearing loss from Hamedan province

Background and aims: Hearing loss is a most common sensory deficit in humans. The hearing loss may be conductive, sensorineural, or mixed (syndromic or nonsyndromic), prelingual or postlingual. Due to the complexity of the hearing mechanism, it is not surprising that several hundred genes might be involved in causing hereditary hearing loss. So far 152 loci have been identified which are associated with the most common type of hearing loss. This study aimed to analyze genetic linkage of DFNB7/11 locus in families with autosomal recessive nonsyndromic hearing loss from Hamedan province. Methods: In this descriptive laboratory study, 24 families from Hamedan province with autosomal recessive nonsyndromic hearing loss were examined. Selected families in this study had consanguinity and they were negative for GJB2 gene mutations. Linkage analysis was performed by 7 markers (STR) for DFNB7/11 locus. Results: After examining different families, one family of the 24 families (4.16%) showed linkage to the DFNB7/11 locus, but in examining different exons and promoter of TMC1 gene, no mutation was found. Lack of TMC1gene mutations in mentioned familyy suggests mutations exist in noncoding regions of TMC1 genes or new gene exists in this locus. SLINK value and LOD score of this family was 1.45 and 0.54 respectively. Conclusion: Based on the results of this study, DFNB7/11 locus may not have important role in causing hearing loss of population studied, but further studies are necessary to determine more precisely the role of this locus in hearing loss in Iranian population

Evaluation of MT1XT20 single quasi-monomorphic mononucleotide marker for characterizing microsatellite instability in persian lynch syndrome patients

Background: Colorectal malignancies with high microsatellite instability (MSI-H), either hereditary (Lynch syndrome) or sporadic, demonstrate better prognosis and altered response to 5FU chemotherapy. It is now recommended to perform MSI testing for all new cases of colorectal cancer regardless of being categorized as hereditary or sporadic. For MSI detection, immunohistochemistry or PCR-based protocols using a cohort of various sets of STR markers are recommended. Here we aimed to evaluate a simplified protocol using just a single STR marker, MT1XT20 mononucleotide repeat, for detection of MSI in Lynch syndrome patients. A Promega five-marker MSI testing panel and immunohistochemistry (IHC) were used as the gold standard in conjunction with MT1XT20. Materials and Methods: Colorectal patients with a positive history of familial cancers were selected by evaluating medical records. Based on Amsterdam II criteria for Lynch syndrome 20 families were short listed. DNA was extracted from formalin fixed paraffin embedded tumour and adjacent normal tissues resected from the index case in each family. Extracted DNA was subjected to MT1XT20 mononucleotide marker analysis and assessment with a commercially available five marker MSI testing kit (Promega, USA). IHC also was performed on tissue sections and the results were compared with PCR based data. Results: Eight (40%), seven (35%) and five (25%) cases were MSI positive using with the Promega kit, IHC and MT1XT20, respectively. Among the markers included in Promega kit, BAT26 marker showed instability in all 8 samples. NR24 and NR21 markers showed instability in 7 (87.5%), and BAT25 and MONO 27 in 6 (75%) and 5 (62.5%). Conclusions: Although MT1XT20 was earlier reported as a valid standalone marker for MSI testing in CRC patients, we could not verify this in our Iranian patients. Instead BAT26 among the markers included in Promega MSI testing kit showed instability in all 8 MSI-H CRC samples. Therefore, it seems BAT26 could act well as a single marker for MSI testing in Iranian CRC patients

Association of (Ile462Val in genetic polymorphisms CYP1A1) and uterine leiomyoma risk in women in Charmahal va Bakhtiari, I.R. Iran

زمینه و هدف: لیومیوم رحمی شایع‌ترین تومور خوش خیم عضلات صاف رحم می‌باشد و تقریباً در ٢٥ تا ٣٠ زنان بالای ٣٠ سال رخ می‌دهد. رشد و پیشرفت لیومیوم وابسته به استروژن بود و نقش ژن سیتوکروم 450P- کلاس 1A1 (CYP1A1) در متابولیسم استروژن بررسی شده است. لذا این مطالعه با هدف بررسی ارتباط پلی مورفیسم Ile462Val (A>G) در ژن CYP1A1 با لیومیوم رحمی در زنان استان چهارمحال و بختیاری انجام شده است. روش بررسی: در این مطالعه مورد - شاهدی ١٥٦ زن با تشخیص کلینیکی لیومیوم رحمی و ١٥١ زن سالم به روش در دسترس انتخاب و مورد مطالعه قرار گرفتند. توزیع ژنتیکی پلی مورفیسم Ile462Val (A>G) ژن CYP1A1 با روش PCR-RFLP تعیین گردید. داده‌ها به کمک آزمون کای دو و t تجزیه و تحلیل شد. یافته‌ها: ژنوتیپ AG در زنان مبتلا به لیومیوم 2/12 (19 نفر) و در گروه شاهد 6/8 (13 نفر) بود. اختلاف معنی‌داری در فراوانی اللی و ژنوتیپی پلی مورفیسم ژن CYP1A1 بین دو گروه یافت نشد (306/0=P). نتیجه‌گیری: نتایج این مطالعه نشان داد که بین پلی مورفیسم ژن CYP1A1 (Ile462Val) و افزایش خطر لیومیوم در جمعیت زنان مورد مطالعه ارتباطی وجود ندار

The frequenc of hearing loss etiology among deaf students in Chaharmahal va Bakhtiari province Iran 2008-2009

چکیده: زمینه و هدف: ناشنوایی شایع ترین اختلال حسی در انسان می باشد که تاثیرات اقتصادی و اجتماعی شدیدی در دنیای مدرن دارد. از نظر اتیولوژی، ناشنوایی به دو دسته ژنتیکی و غیر ژنتیکی تقسیم می شود که ناشنوایی ژنتیکی در دو گروه سندرمیک و غیر سندرمیک طبقه بندی می گردد. این مطالعه با هدف تعیین اتیولوژی ناشنوایی در دانش آموزان ناشنوای استان چهارمحال و بختیاری انجام شد. روش بررسی: در این مطالعه توصیفی 265 دانش آموز ناشنوای خفیف تا عمیق شرکت داشتند. نمونه ها شامل دانش آموزان ناشنوای مدارس استان چهارمحال و بختیاری بودند که دامنه سنی آنها بین 6 تا 22 سال بود. سابقه پزشکی، شجره نامه و اطلاعات دموگرافیک بیماران با استفاده از پرسشنامه جمع آوری گردید. تمام بیماران مورد معاینه عمومی و شنوایی قرار گرفته و تست Pure Tone Audiometry) PTA) برای کلیه دانش آموزان و تست ABR (Auditory Brainstem Response)، OAE(Otoacoustic Emissions) در بیماران مشکوک به ناشنوایی عصبی انجام شد. یافته ها: 2/67 از والدین دانش آموزان ناشنوا ازدواج خویشاوندی داشته و شایع ترین شکل این ازدواج، ازدواج بین خویشاوندان درجه 3 (1/78) بود. نتایج مطالعه نشان داد که 8/98 موارد ناشنوایی ژنتیکی به شکل اتوزمال مغلوب و در 8/97 موارد ناشنوایی از نوع حسی عصبی بود. از نظر اتیولوژی ناشنوایی با علت ژنتیکی، شایع ترین علت ناشنوایی(8/60) و سپس ناشنوایی با علل ناشناخته و اکتسابی بیشترین موارد را تشکیل می دادند. در 2/4 از دانش آموزان علل سندرمیک مطرح و مشکلات چشمی بیشترین ناهنجاری همراه با ناشنوایی بود. در بین علل اکتسابی نیز بیشترین عامل ناشنوایی به ترتیب مربوط به کرنیکتروس و تشنج همراه با تب بود. نتیجه گیری: نتایج این مطالعه اهمیت مشکلات مربوط به ازدواج های خویشاوندی را در جمعیت مورد مطالعه نشان می دهد. میزان این ازدواج ها در جمعیت مورد مطالعه بسیار بالا (2/67) بوده که می تواند علت اصلی ناشنوایی مادرزادی باشد

Genetic study of 45 big hearing loss pedigrees and GJB2 gene mutations frequency in Chaharmahal va Bakhtiari province, Iran, 2008

Background and aim: Hearing loss is the most common sensorineural disorder in human. Despite the contribution of several different genes in causing deafness, mutations in the GJB2 gene have been involved in deafness in many populations. This study aims to investigate genetic epidemiology and frequency of GJB2 gene mutations in 45 big deaf pedigrees. Methods: In this genetic epidemiology study we have investigated 45 big deaf pedigrees concerning inheritance patterns, consanguinity and diversity of deafness severity among siblings using data collected by questionnaires and audiograms. We examined also the frequency and profile of GJB2 gene mutations in 45 probands using direct sequencing strategy. Results: Our study revealed 73% of consanguinity in the deaf pedigrees studied. The most common type of first cousins marriage was found between first cousins who were the children of two brothers. We found autosomal recessive and X-linked recessive pattern in 94-97% and 3-6% of the pedigrees studied respectively. Regarding molecular analysis, GJB2 mutations were found in 11% of population studied including 35delG, 167delT, 299-300delAT and 363delC. Conclusion: A high rate of consanguineous marriage determined in this study could raise the rate of autosomal recessive patterns up to 94-97% of the overall pedigree and would be the main cause to prepare the way for congenital deafness. This study revealed a low contribution of GJB2 gene mutations in causing deafness in Chaharmahal va Bakhtiari province