[Detection and typing of human papilloma virus by polymerase chain reaction and hybridization assay in cervical samples with cytological abnormalities]. / Sitolojik atipi izlenen servikal örneklerde insan papilloma virusunun polimeraz zincir reaksiyonu ve hibridizasyon yöntemleriyle saptanmasi ve tiplendirilmesi.

Certain mucosa-tropic human papillomavirus (HPV) types are associated with carcinoma of the uterine cervix or its precursor lesions. In addition to cytological screening, early diagnosis and treatment of cervical carcinoma rely on sensitive detection and typing of HPV isolates. In this study, HPV detection and typing were performed in the cervical samples of patients with abnormal cytological evaluation. Forty randomly-selected cervical samples that comprise 18 ASC-US (Aypical Squamous Cells of Undetermined Significance), four AG-US (Aypical Glandular cells of Undetermined Significance), one ASC-H (Atypical Squamous Cells-can not exclude HSIL), one HSIL (High-grade Intraepithelial Lesion), 14 LSIL (Low-grade Intraepithelial Lesion), one adenocarcinoma and one squamous cell carcinoma, obtained by a commercial liquid-based cytology system (ThinPrep Pap Smear Method, Cytyc, USA), were included to the study. HPV-DNA detection were accomplished by L1 in-house polymerase chain reaction (PCR) performed using MY09/11 and GP5/6 primers along with a commercial real-time PCR (HeliosisTM HPV LC PCR Kit; Metis Biotechnology, Turkey) that detects HPV infections and HPV-16 via melting curve analysis. A commercial PCR-array hybridization test (Rapid HPV Genotyping MacroArray; HybriBio Inc, Hong Kong) that can identify 21 low and high risk HPV types was employed for typing. Viral DNA was detected in 35% (14/40) and 57.5% (23/40) of the samples by MY09/11 and GP5/6 primers, respectively. All in-house PCR positive samples were also positive in the real-time PCR assay. PCR-array hybridization assay provided typing results in 95.6% (22/23) of the PCR positive samples while one LSIL sample could not be typed by any of the methods used. High risk HPV types 16, 18, 31, 45, 52, 56, 58, 59,68 (65.8%); probable high risk type 53 (13.2%), low risk types 6, 42 and 81 (21%) were identified out of a total of 38 HPV isolates. Multiple infections with more than one HPV type were identified in 45.5% (10/22) of positive samples. High/probable high risk types were detected in all single infections and all low risk isolates were present in multiple infections. HPV-16 was identified in 31.8% (7/22) by real-time PCR and in 45.5% (10/23) of positive samples by PCR-array hybridization assay. HPV-16 was observed to be the most frequently detected type (10/22, 45.5%), followed by types 53 and 81 (5/22, 22.7%); 68 (4/22, 18.2%); type 58 (3/22; 13.6%); types 31, 42 and 59 (2/22; 9.1%) and others (1/22, 4.5%). As a result our data have indicated the abundance of high risk HPV isolates and infections with multiple HPV types in that specific area.

Renin-angiotensin system expression in the K562 human erythroleukaemic cell line.

Local renin-angiotensin system (RAS) may affect leukaemic cell production within the bone marrow microenvironment. Angiotensin-converting enzyme (ACE), renin, and angiotensin could influence leukaemogenesis. In this study, mRNA expressions of the major RAS components (ACE, renin, and angiotensinogen) in K562 human erythroleukaemia cell line have been searched by Real Time quantitative polymerase chain reaction. K562 blasts are multipotential, haematopoietic malignant cells that spontaneously differentiate into recognisable progenitors of the erythrocyte, granulocyte and monocytic series. We observed significant expressions of ACE, renin, and angiotensinogen in K562 leukaemic blast cells. Therefore, K562 human erythroleukaemia cell line may serve as an in vitro model to elucidate the role of RAS in leukaemia and to test the effects of RAS-affecting drugs on leukaemic cellular proliferation.

Alterations in the 'local umbilical cord blood renin-angiotensin system' during pre-eclampsia.

BACKGROUND: Local autocrine-paracrine renin-angiotensin system (RAS), independently functioning from the circulating RAS, is present in major organs of the female reproductive tract. We have previously demonstrated 'a local RAS in human umbilical cord' via verifying the corresponding ACE, renin, and angiotensinogen mRNAs. The aim of this study is to search alterations of the local umbilical cord RAS during pre-eclampsia. METHODS: Cord blood samples were obtained from 19 patients with pre-eclampsia (aged mean 26.6 ± 5.83 (range 18-42) years) and 20 women with normal pregnancy (aged mean 28.26 ± 7.30 (range 19-37) years). Women with uncomplicated pregnancy formed the control group. Real time quantitative PCR analysis for ACE, renin and angiotensinogen gene expressions were carried out using a LightCycler™ instrument. RESULTS: The mean expression ratios were 0.0029 ± 0.0015 for renin, 0.153 ± 0.166 for angiotensinogen, and 0.220 ± 0.294 for ACE, in control samples. The mean expression ratios of pre-eclamptic patients were 0.0061 ± 0.00068, 0.035 ± 0.008, and 0.030 ± 0.006 for renin, angiotensinogen and ACE genes, respectively. While renin expressions increased in the local cord blood of pre-eclampsia in comparison to the normal cord blood, unpredictable decrements in the angiotensinogen and ACE expressions were observed within the same pre-eclamptic samples. There were no statistically significant differences between intrauterine growth restriction (IUGR) and appropriate for gestational age (AGA) newborns in respect to renin, angiotensinogen and ACE gene expressions. CONCLUSIONS: These findings indicate that the gene expression in the major components of the local RAS does not represent a constant mathematical model, but is affected from the ongoing pathobiological events associated with the disease course. Local umbilical cord blood RAS alterations at the basis of genetic expression are evident in pre-eclampsia.

[Human papilloma virus DNA in cervical samples with cytological abnormalities and typing of the virus]. / Sitolojik olarak anomali saptanan serviks örneklerinde insan papilloma virus DNA'sinin arastirilmasi ve virusun tiplendirilmesi.

Human Papillomavirus (HPV) infections and cervical carcinoma which is associated with certain types of the virus have been worldwide public health issue. Early diagnosis of HPV infections and the detection of viral genotypes are important for the successful treatment and prevention of cervical carcinoma. This study has been designed as a preliminary study to estimate HPV type distribution in cervical samples with cytologic abnormalities in our country. A total of 35 cervical samples which were evaluated by a commercial liquid-based cytological system, were included to the study. The presence of HPV-DNA has been searched with nested polymerase chain reaction (PCR) by using consensus primer sets of MY09/11 and GP5/6 that target L1 region of the viral genome. HPV typing was performed by direct sequencing of the amplicons. In cytologic evaluation, 14 samples were diagnosed as ASC-US (Aypical squamous cells of undetermined significance), three were ASC-H (Atypical squamous cells-cannot exclude HSIL), five were HSIL (High-grade intraepithelial lesion), seven were LSIL (Low-grade intraepithelial lesion), four were LSIL+suspected HSIL, one was AG-US (Aypical glandular cells of undetermined significance) and one was atypical cells of undefined nature. HPV-DNA was detected in 28 of the 35 (80%) samples, and sequence analysis revealed high-risk HPV types (type 16, 18, 31, 33, 45, 56, 59) in 22 (78.6%) samples, probable high-risk types (type 53) in two (7.1%) samples and low-risk types (type 6, 54, 72, 81) in four (14.3%) samples. HPV type 16 emerged as the most frequently-detected type, comprising 50% (14/18) of all samples; followed by type 18 in 10.7% (3/28) and type 53 in 7.1% (2/28) of the samples. As a result, although the number of cervical samples were relatively low, the preliminary data obtained with this study revealed the HPV type distribution, however more detailed studies are needed to elucidate the epidemiology of HPV infections in Turkey.

Effect of erythropoietin on brain tissue after experimental head trauma in rats.

BACKGROUND: The purpose of this study was to investigate the effect of EPO on LPO, on ultrastructural findings, and on antiapoptotic bcl-2 and survivin gene expressions after TBI. The authors also compared the activity of EPO with that of MPSS. METHODS: Wistar rats were divided into 6 groups: sham-operated, control, moderate TBI-alone (300 g/cm), TBI + EPO-treated (1000 IU/kg), TBI + MPSS-treated (30 mg/kg), and TBI + vehicle-treated (0.4 mL albumin solution) groups. RESULTS: Compared with the levels in control and sham-operated animals, LPO was significantly elevated in rats in the trauma-alone group. The administration of EPO and MPSS significantly decreased the LPO levels (P < .05). Trauma also increases the antiapoptotic bcl-2 gene expression significantly at 24 hours postinjury (P < .05), but it has no effect on survivin expression. The EPO and MPSS treatments caused significant elevation in both gene expressions (P < .05). It is also showed that MPSS has more protective effect than EPO on brain ultrastructure, especially on the structure of small- (P < .05) and medium-sized myelinated axons, after TBI. CONCLUSIONS: EPO has protective effects after moderate TBI, and this effect seems better than MPSS on antiapoptotic gene expression and LPO. The protection of cerebral subcellular organelles after traumatic injury is more prominent in MPSS-treated animals than EPO-treated animals quantitatively. This experimental study indicates that the benefits of EPO in the management of TBI have promising results and prompts further studies on the difference between EPO and MPSS in histopathological findings at the subcellular level.

Overexpression of the local bone marrow renin-angiotensin system in acute myeloid leukemia.

OBJECTIVES: Local bone marrow renin-angiotensin system (RAS) is an autocrine-paracrine system affecting hematopoiesis. Angiotensin II stimulates the proliferation of bone marrow and umbilical cord blood hematopoietic progenitors. Angiotensin-converting enzyme (ACE) hyperfunction may lead to the acceleration of negative hematopoietic regulator peptide, AcSDKP, metabolism, which in turn lowers its level in the bone marrow microenvironment, finally removing the antiproliferative effect of AcSDKP on the hematopoietic cells and blasts. The aim of this study is therefore to search those major RAS components simultaneously in the leukemic blast cells taken from the bone marrow of patients with acute myeloid leukemia (AML). METHODS: Bone marrow aspiration materials were obtained from 10 patients with AML (8 males, 2 females; median age 48.5 years) and 8 patients with nonmalignant hematological disorders (6 males, 2 females; median age 45 years). EDTA-treated bone marrow samples were stored at -70 degrees C until analysis. Total RNA was extracted from 200-microl bone marrow samples by High Pure RNA Isolation Kit. RESULTS: The medians of expression ratios of AML patient samples have been found 0.736 (IQR 1.359), 0.540 (IQR 0.725), and 0.075 (IQR 0.002) for ACE, ANG and REN genes, respectively. All three gene expressions were found to be significantly higher in the bone marrow samples of AML patients. CONCLUSION: In this study, the expression of the mRNAs of the major RAS components-namely ACE, renin and angiotensinogen-in human bone marrow samples were quantified by reverse transcription-polymerase chain reaction (RT-PCR) to confirm the presence of the local bone marrow RAS. Elucidation of the pathological activity of the local RAS-mediated regulation of the leukemogenesis is both pathobiologically and clinically important, since the angiotensin peptides represent a molecular target in the disease management.

Developmental potential of bovine oocytes cultured in different maturation and culture conditions.

Diverse groups of chemicals in culture media are needed for successful bovine oocyte maturation and embryo development during which dramatic cytoplasmic and nuclear reprogramming events take place. In vitro embryo production (IVP) procedures frequently include supplements such as serum and/or co-culture with various types of somatic cells. However, the presence of undefined serum in culture media introduces a variation from batch to batch, increases viral or prion contamination risk, and leads to problems during fetal development. The aim of the present study was to investigate the possibility of using chemically defined-synthetic serum substitute (SSS) in place of fetal calf serum (FCS) during maturation and long-term culture to stimulate in vitro maturation (IVM), fertilization (IVF) and subsequent embryo development. In Experiment I, the effect of the protein source on in vitro maturation was tested by maturing oocytes in culture media supplemented with 10% FCS (Control Group), 10% SSS (Group I) and 10% SSS+10 ng/ml epidermal growth factor (EGF) (Group II). In Experiment II, effects of SSS on both oocyte maturation and embryo development during in vitro culture (IVC) were tested by maturing oocytes in media supplemented with 10% FCS (FCS Group) or 10% SSS+10 ng/ml EGF (SSS Group), followed by IVF and IVC in SOF media supplemented with 10% FCS and 10% SSS on day 4 for FCS and SSS Groups, respectively. Even though rates for cleavage and development to blastocyst stage were not different, blastocyst cell numbers were higher in Group II containing SSS and EGF. The SSS supplementation group had higher apoptotic nuclei as compared to the FCS Group in Experiment II. Transcripts for heat shock protein 70 (Hsp70), interferon tau (IF-tau), DNA methyltransferase 3a (Dnmt3a), desmosomal glycoprotein desmocollin III (DcIII) and insulin-like growth factor II receptor (Igf-2r) were altered in different culture conditions in Experiment I. However, only glucose transporter-1 (Glut-1) mRNA was different in the SSS and FCS Groups in the second experiment. In summary, SSS and EGF in maturation medium and replacement of FCS with SSS alone in culture medium on day 4 of IVC support oocyte maturation and embryo development in vitro. However, significance of culture condition induced changes on the genome-wide abundance of messenger ribonucleic acid and the significance of the apoptotic nuclei during fetal development still remain to be determined.

Prevalence of Helicobacter pylori in symptomatic patients and detection of clarithromycin resistance using melting curve analysis.

UNLABELLED: Abstract. BACKGROUND: Clarithromycin is often a component of combination therapies for Helicobacter pylori eradication; however, increases in resistance rates have decreased the success of the treatment. OBJECTIVE: This study was designed to determine the prevalence of H pylori infection in symptomatic patients and to detect clarithromycin resistance rates using melting curve analysis. METHODS: Patients scheduled for upper endoscopy at the Endoscopy Unit of the Department of Gastroenterology, Duzce University, Medical Faculty Hospital, Konuralp/Duzce, Turkey, were assessed for enrollment in the study. Two pairs of gastric biopsy specimens (antrum and corpus) were obtained from each study patient. Histopathologic examination, rapid urease test, culture, and polymerase chain reaction (PCR) of the specimens were used to identify H pylori infection. Clarithromycin resistance was detected using melting curve analysis. RESULTS: Seventy-five patients (41 women, 34 men; mean [SD]age, 42.6 [14.5] years [range, 17-70 years]) were included in the study. Using histopathology and rapid urease test, H pylori was detected in 40 (53.3%) of the 75 specimens. H pylori was detected using PCR in 40 (53.3%) specimens and by culture in 10 (13.3%) specimens. The specificity and sensitivity of PCR and culture were interpreted by comparing them with the results of histopathologic examination and urease tests. The specificity and sensitivity of PCR were 68.6% and 72.5%, respectively, and the specificity and sensitivity of culture were 97.1% and 22.5%, respectively. Of the 40 isolates, 21 (52.5%) were susceptible to clarithromycin, 12 (30.0%) were resistant, and a mixed susceptibility pattern was detected in 7 (17.5%) specimens. H pylori isolates from 19 (79.2%) of the 24 patients who had formerly used clarithromycin showed clarithromycin resistance. CONCLUSIONS: The prevalence of H pylori infection was 53.3% for the symptomatic patients in this study, and 47.5% of the isolates showed clarithromycin resistance using melting curve analysis. The PCR-based system used in this study was accurate for the detection of H pylori infection as well as clarithromycin susceptibility testing directly in biopsy specimens.

Expression of cyclooxygenase-2 in orbital fibroadipose connective tissues of Graves' ophthalmopathy patients.

OBJECTIVE: The aim of this study is to evaluate the expression of cycloocygenase-2 (COX-2) in orbital fibroadipose connective tissue in Graves' ophthalmopathy (GO) patients, and investigate the associations between COX-2 expression and GO characteristics. METHODS: The orbital fibroadipose connective tissues of 23 cases demonstrating moderate or severe GO, and eight control subjects without any history of thyroid or autoimmune disease were analyzed for COX-2 mRNA expression. Real-time relative quantitative PCR was performed to assess transcripts of COX-2 using the LightCycler. The disease activity was evaluated by the clinical activity score (CAS). The clinical features of GO were evaluated by total eye score (TES) and the cases were divided into two groups; type 1 cases included higher degrees of proptosis with orbital fat volume increase, and type 2 cases included cases with compressive neuropathy and limited extraocular muscle functions. RESULTS: The mean +/- s.d. disease duration was 5.7 +/- 7.1 years. The mean +/- s.d. CAS and TES of cases were 1.60 +/- 1.04 and 7.5 +/- 1.8 respectively. The mean +/- s.d. expression of COX-2 was 0.023 +/- 0.013 and 0.010 +/- 0.002 in GO cases and controls (P = 0.008), and 0.015 +/- 0.073 and 0.029 +/- 0.135 in type 1 and type 2 cases respectively (P = 0.007). COX-2 expression showed a statistically significant positive correlation with TES (r = 0.634, P = 0.001), and a negative correlation with the disease duration (r = -0.621, P = 0.002). CONCLUSIONS: COX-2 is expressed at higher levels in orbital fibroadipose tissues of GO cases. This showed a positive correlation with increasing severity of orbital disease suggesting possible relation with COX-2 expression and orbital inflammation in GO.

Developmental and molecular correlates of bovine preimplantation embryos.

Expression of embryonic genes is altered in different culture conditions, which influence developmental potential both during preimplantation and fetal development. The objective of this study was to define the effects of culture conditions on: bovine embryonic development to blastocyst stage, blastocyst cell number, apoptosis and expression patterns of a panel of developmentally important genes. Bovine embryos were cultured in vitro in three culture media containing amino acids, namely potassium simplex optimization medium (KSOMaa), Charles Rosenkrans 1 (CR1aa) and synthetic oviductal fluid (SOFaa). Apoptosis in blastocysts was determined by TUNEL assay and expression profiles of developmentally important genes were assayed by real-time PCR. In vivo-produced bovine blastocysts were used as controls for experiments determining gene expression patterns. While the cleavage rates did not differ, embryos cultured in SOFaa had higher rates of development to blastocyst stage (P < 0.05). Mean cell numbers and percentages of apoptotic cells per blastocyst did not differ among the groups. Expression of the heat shock protein 70 (Hsp70) gene was significantly up-regulated in both CR1aa and KSOMaa when compared with SOFaa (P < 0.001). DNA methyltransferase 3a (Dnmt3a) expression was higher in embryos cultured in CR1aa than in those cultured in SOFaa (P < 0.001). Expression of interferon tau (IF-tau) and insulin-like growth factor II receptor (Igf-2r) genes was significantly up-regulated in KSOMaa when compared with CR1aa (P < 0.001). Gene expression did not differ between in vivo-derived blastocysts and their in vitro-derived counterparts. In conclusion, SOFaa supports higher development to blastocyst stage than KSOMaa and CR1aa, and the culture conditions influence gene expression.

Renin-angiotensin system gene polymorphisms: association with susceptibility to Henoch-Schonlein purpura and renal involvement.

The clinical course of Henoch-Schönlein Purpura (HSP) in children is variable, with some patients having a much more rapidly progressing course than others. We investigated whether polymorphisms of the renin-angiotensin system (RAS) genes are involved in HSP. Three RAS genotypes were examined in 114 children with HSP and in 164 healthy children: the angiotensin I converting enzyme (ACE) insertion/deletion polymorphism, the M235T mutation in the angiotensinogen gene (Agt), and the A1166C in the angiotensin II type I receptor (AT1R) gene. Significant differences were observed between HSP patients and control group in the frequency of ACE and Agt genotypes (p=0.004 and p=0.003, respectively). The TT genotype of Agt gene was associated with a 3.5-fold increased risk for Henoch-Schönlein nephritis (HSN) compared with the MM/MT genotype (odds ratio, 3.5; 95% confidence interval, 1.2-10.4). There was a trend to a higher prevalence of the TT genotype of the Agt gene among patients with nephrotic range proteinuria when compared to the patients with mild proteinuria, although the difference did not reach a statistical significance. The results of this study suggest that polymorphisms of ACE gene and Agt gene likely influence the risk of developing HSP. However, among the three genes of the RAS studies, only Agt gene was associated with the susceptibility to HSN. RAS gene polymorphisms studied are not associated with the presence of nephrotic range proteinuria. Additional studies are warranted to verify the correlation between RAS gene polymorphisms and susceptibility to HSP.

Apoptosis-related gene bcl-2 in lung tissue after experimental traumatic brain injury in rats.

BACKGROUND: We have recently shown that experimental traumatic brain injury resulted in ultra structural damage in lung tissue. The main objective of the current study was to investigate in a rat model of brain injury whether expression of Bcl-2 gene and lipid peroxidation levels in the lung tissue after traumatic brain injury were affected by methylprednisolone sodium succinate (MPSS) treatment. METHODS: Fifty-six Wistar-Albino female rats weighing 180-220 g were used, which were allocated into seven groups. A weight-drop method was used to achieve head trauma. Real time quantitative PCR analyses for Bcl-2 gene expression and measurement of the levels of lipid peroxidation were carried out. All the data was analyzed by using SPSS 11.5 for Windows. RESULTS: Mean Bcl-2 expression in the methylprednisolone group was considerably higher compared to that of all the other groups (p<.05). Mean lipid peroxidation levels were significantly higher in the trauma group and notably lower in the methylprednisolone group (p<.01). CONCLUSIONS: The oxidative stress imposed on lung tissue, as seen by high levels of lipid peroxidation, after brain injury was significantly attenuated by MPSS treatment. MPSS treatment following brain injury also augmented putative anti-apoptotic Bcl-2 gene expression in lung tissue. Further studies are required to determine the full range and lower limits of effective MPSS dose. More importantly the optimal efficacy according to the timing of MPSS treatment after brain injury needs to be determined for impact on more diverse markers of cell inflammation, apoptosis and injury.

Gene polymorphisms in patients below 35 years of age who underwent coronary artery bypass surgery.

OBJECTIVE: Genetic bases for novel prothrombotic, inflammatory risk factors may play a role in the early onset of coronary artery disease. METHODS: Twenty-one patients below 35 years of age who underwent coronary bypass grafting between 2002 and 2004 constituted the study group and were compared with 50 healthy, age and sex-matched controls. Gene analysis for genetic polymorphisms of angiotensin-converting enzyme, prothrombin G20210A, tumour necrosis factor-alpha G308A, factor V Leiden and interleukin-6 genes was carried out. RESULTS: The control group was 98% homozygous for the factor V Leiden GG allele and 2% heterozygous for the GA allele. On the other hand, the study group was 76.2% homozygous for the GG allele, and 23.8% heterozygous for the GA allele (P<0.05). Homozygosity for factor V Leiden mutation (AA) was not encountered in either group. With regard to interleukin-6, 70.0% of the control group demonstrated homozygosity for the GG allele and 30.0% showed heterozygosity (GC). The study group was 52.4% homozygous for the GG allele and heterogenicity was similar in this group (28.6% GC). On the other hand, 19.0% of this group demonstrated CC homogenicity (P<0.05). No difference was observed with regard to gene polymorphisms. CONCLUSIONS: Gene polymorphisms with regard to prothrombotic factor V Leiden mutation and inflammatory marker interleukin-6 may play a role in the pathogenesis of early-onset coronary artery stenosis in patients below 35 years of age.

Relationship between HLA-DR, HLA-DQ alleles and MEFV gene mutations in familial Mediterranean fever (FMF) patients.

BACKGROUND/AIMS: Three missense mutations clustered on the carboxyl-terminal portion of the MEFV gene (M680I, M694V, and V726A) have been observed in over 80% of affected alleles in several ethnic groups of familial Mediterranean fever patients. Several immunologic abnormalities were found both in cellular and humoral components in Mediterranean fever patients. Those observations have pointed the way for analysis of the HLA region in Mediterranean fever. We intended to compare HLA DR/DQ alleles with those major mutations in the MEFV gene in Mediterranean fever patients. METHODS: The distribution of MEFV gene mutations and HLA-DR, HLA-DQ alleles were analyzed in 40 index Turkish Mediterranean fever patients, 28 family members and 42 healthy controls. M680I, M694V, and V726A mutations were studied by amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) analysis. HLA-DR and DQ allele subgroups were studied using SSP-PCR technique. RESULTS: A total of 37 (92.5%) patients in 40 Mediterranean fever index patients were found to carry one of the three missense mutations. The HLA-DR4 allele frequency was significantly higher in the Mediterranean fever patient group. When comparisons were made between Mediterranean fever mutations and HLA allele frequencies, M694V mutation with HLA DR3, DR11/5 and DR 13/6 and M680I mutation with DR7 allele subgroups were statistically significant. DQ6/1, DQ7/3, and DQ8/3 allele with M694V, DQ2 allele with M680I, and DQ6/1 with V726A mutations were also statistically significant. CONCLUSIONS: Our results indicate a relationship between some HLA-DR/DQ alleles and MEFV mutations in Mediterranean fever patients. We suggest HLA-DR/DQ alleles and their role in the pathogenesis of Mediterranean fever need further analysis and comparative studies.

Renin-angiotensin system polymorphisms and coronary artery surgery patients.

The frequencies of angiotensin-converting enzyme gene insertion/deletion, angiotensinogen-M253T, and angiotensin II type 1 receptor-A1166C polymorphisms were analyzed in 105 patients undergoing coronary artery bypass grafting (group 1) and a control group of 105 non-cardiac patients (group 2). Blood samples were obtained for biochemical analyses and DNA extraction. Genotyping was performed by polymerase-chain-reaction-based restriction analysis. According to the angiotensin-converting enzyme gene insertion/deletion polymorphism, 36.3% of patients in group 1 and 30.7% in group 2 were homozygous for the DD allele. This difference was not statistically significant. Angiotensin II type 1 receptor-A1166C genotype polymorphism was also not significantly different between the groups. The results showed the angiotensinogen-M235T polymorphism to be heterogenous. The MM homozygote frequency was significantly higher in controls (72.3%), whereas 80% of the TT homozygote frequency was in the surgical group ( p = 0.001). These results show that although there were no significant differences in angiotensin-converting enzyme gene insertion/deletion and angiotensin II type 1 receptor-A1166C genotype polymorphisms between the groups, angiotensinogen-M235T polymorphism of TT homozygote frequency was significantly associated with patients undergoing coronary artery bypass surgery.

Local umbilical cord blood renin-angiotensin system.

Local bone marrow (BM) renin-angiotensin system (RAS) is an autocrine-paracrine system affecting normal and neoplastic hematopoiesis. Angiotensin II type 1a (AT1a) receptors are present on the CD34(+) hematopoietic stem cells (HSC). Angiotensin II stimulates the proliferation and differentiation of the HSC populations through the activation of AT1 receptors on HSC. Umbilical cord blood (UCB) is a rich source of HSC. The existence of a complete local UCB RAS has not been previously investigated. In this study, local synthesis of the major RAS components, namely, angiotensin-converting enzyme (ACE), renin, and angiotensinogen, was identified by demonstrating their corresponding mRNAs using quantitative reverse transcriptase polymerase chain reaction (RT-PCR) in human UCB. Local RAS could regulate cellular growth in a variety of tissues including the BM. Major RAS peptides can exert significant effects on primitive pluripotential HSC populations. Further studies should focus on the interactions between possible autocrine, paracrine, endocrine, and intracrine actions of the local UCB RAS and growth, engraftment, differentiation, and plasticity functions of HSC of UCB origin.

Vitamin D receptor gene polymorphism in hypercalciuric children.

Idiopathic hypercalciuria is a complex disease resulting from an interaction between environmental and genetic factors. Recently, the relationship between vitamin D receptor ( VDR) alleles and calcium homeostasis has been investigated. This study was conducted to explore the association of VDR gene polymorphism with the risk of absorptive hypercalciuria (AH). We investigated the VDR gene polymorphisms, ApaI, BsmI, and TaqI, in relation to intact parathormone (PTH), osteocalcin, and 25-hydroxyvitamin D in 80 children (42 males, 38 girls) with AH and in 86 healthy children without hypercalciuria. A significant difference in the ApaI genotype was observed between the AH group and the control group ( chi(2)=7.21, P=0.027). The AA genotype was associated with a 3.5-fold increased risk for idiopathic hypercalciuria compared with the Aa/aa genotype (odds ratio 3.5, 95% confidence interval 1.1-11). The BsmI and TaqI polymorphisms did not show any significant association with AH. Serum osteocalcin levels were significantly higher in the group with the AA genotype compared with those with the Aa or aa genotype ( P=0.02, P=0.05, respectively). The results indicate that the ApaI AA genotype of the VDR gene is not only associated with AH but is also related to differences in serum osteocalcin.

The renin-angiotensin system genetic polymorphisms and rheumatic mitral valve disease.

BACKGROUND AND AIM OF THE STUDY: Angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism, angiotensinogen (AGT) gene polymorphism and angiotensin II type 1 receptor (AT1R) polymorphism in relation to rheumatic mitral valve disease were examined in a case-control study to investigate possible relationships between these gene polymorphisms and rheumatic mitral valve disease in patients undergoing mitral valve replacement (MVR). METHODS: A total of 50 patients with rheumatic mitral valve disease and undergoing MVR was compared with 50 normal, and age- and sex-matched control subjects. ACE I/D, AGT gene M235T and AT1R-adenine/cytosine 1166 (A1166C) genotype polymorphisms were identified by polymerase chain reaction (PCR) -based restriction analysis. RESULTS: ACE I/D polymorphism differed significantly between the groups. The control group mostly represented the heterozygote ID allele (74%), while the MVR group showed frequencies of 60% for the homozygote DD and II alleles. MM homozygote frequency was significantly greater in controls, but TT homozygote frequency was significantly greater in the MVR group. AT1R-A1166C genotype polymorphism also differed significantly between groups; the MVR group had 73.7% of the AC heterozygote allele, while controls had 64.4% of the AA and 66.7% of the CC homozygote alleles. CONCLUSION: These results provided evidence of an association between ACE I/D polymorphism, M235T polymorphism and AT1R-A1166C genotype polymorphism and rheumatic mitral valve disease.

Polymorphisms in the vitamin D receptor gene and the risk of calcium nephrolithiasis in children.

OBJECTIVE: Polymorphism in the Vitamin D Receptor (VDR) gene has recently been reported to be associated with calcium metabolism disorders. This study was conducted to investigate the association of VDR gene polymorphism with the risk of calcium nephrolithiasis. METHODS: We investigated the VDR ApaI, BsmI and TaqI polymorphisms, in relation to serum calcium, phosphate, intact parathyroid hormone and 1.25(OH)(2)D(3) in 64 hypercalciuric stone-forming children and 90 healthy children. DNA was isolated from peripheral blood, and genotyping was performed with PCR-based methods. RESULTS: The frequency of ApaI AA genotype was significantly higher in the children with calcium nephrolithiasis than the controls (chi(2)=9.5; p=0.008). The distribution of BsmI and TaqI genotypes in stone-forming patients was similar to those in the control group. There was a significant association between TaqI TT genotype and the strength of the family history. The patients with TT genotype were observed to have a 8 times more risk than patients with Tt/tt genotype for recurrent stone episodes (OR 8, 95%CI 1.61-39.6). CONCLUSION: VDR genotype determination may provide a tool to identify individuals who are at a risk for calcium nephrolithiasis.