Post-transcriptional gene silencing: Basic concepts and applications

Post-transcriptional gene silencing (PTGS)-mediated gene silencing exploits the cellular mechanism whereintranscripts having sequence similarity to the double-stranded RNA (dsRNA) molecules present in the cell will besubjected to degradation. PTGS is closely related to natural processes such as RNA-mediated virus resistance andcross-protection in plants. Gene silencing and the cellular machinery for affecting this phenomenon might haveevolved as a natural protective measure against viral infection in plants. In PTGS, small interfering RNA (siRNA)molecules of 21–23 nucleotides length act as homology guides for triggering the systemic degradation of transcriptshomologous to the siRNA molecules. PTGS phenomenon, first discovered in transgenic petunia plants harbouringchalcone synthase gene and termed co-suppression, has been subsequently exploited to target specific gene transcripts for degradation leading to manifestation of desirable traits in crop plants. Targeted gene silencing has beenachieved either through the introduction of DNA constructs encoding dsRNA or antisense RNA or by deploying cosuppression constructs producing siRNAs against the transcript of interest. Understanding the mechanism of genesilencing has led to the development of several alternative strategies for inducing gene silencing in a precise andcontrolled way. This has paved the way for using PTGS as one ofthe chief functional genomicstools in plants and hashelped in unraveling the mechanism of many cellular processes and identifying the focal points in pathways, besides,opening new vistas in genetic engineering of plants for human benefits. PTGS has shown great potential in silencingthe deleterious genes efficiently so that value-added plant products could be obtained. Thus, PTGS has ushered in anew era in the genetic manipulation of plants for both applied and basic studies. In this review, we have outlined thebasics of RNAi-mediated gene silencing and summarized the work carried out at our institute using this approach, ascase studies. In particular, adopting RNAi-mediated gene silencing (a) as a method to restore fertility in transgenicmale sterile lines developed based on orfH522 gene from sunflower PET1-CMS source, (b) as a tool to suppress theproduction of toxic proteins, ricin and RCA, in castor, and (c) as an approach to induce bud necrosis virus resistancein sunflower has been discussed. Examples from other plant systems also have been mentioned to exemplify theconcept and utility of gene silencing in crop plants.

Role of vitamin D in prevention of adverse maternal and perinatal outcome: a randomized controlled trial

Background: Vitamin  D  deficiency  is  thought  to  be   common among  pregnant  women  and  is  associated  with  adverse  maternal  and  perinatal  outcome. Maternal  and foetal  outcome  in  pregnant  women  with  standard  obstetric  care  was compared with   women with  additional  vitamin D  supplementation.Methods: A randomized comparative  study  was  conducted  on  100  patients attending  the  antenatal  clinic at JSS Medical College and Hospital, Mysuru, Karnataka, India   who were randomly  grouped   into  group A (50 patients) who  received  standard  obstetric care  (500 mg calcium+200 IU vitamin D)  and  group  B (50 patients)  who  received  in  addition  to  standard  obstetric  care   supplementation  of   Vitamin  D  1000 IU/day starting  from  14  weeks  of  gestation  till  delivery.  Vitamin  D  levels  were assessed  in  both  the  groups  with  onset  of  labour  by  chemiluminescence immunoassay  and  obstetric  and  neonatal  outcomes  in  both  groups  were compared.Results: High  incidence  of vitamin  D  deficiency  (96%)  in  standard  care  group  compared  to  vitamin  D supplemented  group  ( p= <0.0001)  was  noted. The  study  showed  significant  reduction  in  risk  of  Preeclampsia (P=0.004),  GDM  (P= 0.02)  and  primary  caesarean  delivery  (0.008)  in  Vitamin  D  supplemented  group. Significantly  high  birth  weight  in  vitamin  D supplemented   group,  an  increase  in  320 grams  in  birth  weight  was  noted  (P <0.0001).Conclusions: There is a high incidence of subnormal vitamin D levels in antenatal women and is associated with maternal and neonatal adverse effects. Measuring Vitamin D levels and appropriate supplementation of higher dose of vitamin D is an effective strategy in prevention of adverse maternal and neonatal outcomes.

Role of diagnostic hysterolaparoscopy in the evaluation of female infertility

Background: Infertility is defined by WHO as a disease of the reproductive system defined by the failure to achieve a clinical pregnancy after 12 months or more of regular unprotected sexual intercourse. The prevalence of infertility is about 10%-15% of reproductive age couples. Female factor is responsible for 40-45% of etiology of infertility. Aim of the study was to evaluate the role of diagnostic hysterolaparoscopy in female infertility at a tertiary care centre.Methods: This is a retrospective hospital based study done at a tertiary care hospital attached to JSS Medical College, Mysuru between January 2017 and December 2018. Infertile women with primary or secondary infertility in the 18-40 years age group, with normal hormonal profile and no known male factor were included.Results: In our study, primary infertility was found in 77% of the 96 patients and secondary infertility was found in 23% of patients. In primary infertility, ovarian pathology was the most common finding while Endometriosis was the most common finding in secondary infertility group. 77 % patients were found to have bilaterally patent tubes while remaining had unilateral or bilateral blockage. On hysteroscopy, endometrial polyps were the commonest hysteroscopic finding in both groups.Conclusions: Combined hystero-laparoscopy is a safe, effective and reliable method in comprehensive evaluation of infertility. It helps in the diagnosis of pelvic pathology which may have been missed by routine examination and thereby helps in optimal management of female infertility.

Maternal pregnancy associated plasma protein-A (PAPP-A) and uterine artery Doppler changes as predictors of pre-eclampsia: a prospective observational study from a teaching hospital in Mysore, Karnataka, India

Background: Hypertensive disorder affects 10-12% of pregnancies. Identifying women, who are at risk is conducive to prompt gestational management. PAPP-A is a protein complex produced by the developing trophoblasts. Low levels of PAPP-A at 10–14 weeks is a marker of impaired placentation and a smaller placental mass. Doppler imaging permits non-invasive evaluation of the uteroplacental circulation and is invaluable in the management of high-risk pregnancies. The uterine artery Doppler screening identifies patients at risk for developing preeclampsia. To study the association of PAPP-A and the uterine artery Doppler changes as predictor of pre-eclampsia in pregnant women at 11-14 weeks of gestation.Methods: This was a prospective study of 150 pregnant women presenting at 11-14 weeks of gestation for a prenatal check-up. After considering the inclusion and exclusion criteria, serum samples for PAPP-A were assayed. Ultrasound Doppler was used to obtain uterine artery flow velocity waveforms and mean pulsatility index and resistance index of uterine arteries were calculated. Cases were followed up till term and observed for development of pre-eclampsia.Results: 48.6% had low serum PAPP-A levels, in which 77% developed PE. The Mean PI and RI is 2.34±1.16 and 0.58±0.1 respectively. 30% women with abnormal PI values and 24% of women with abnormal RI values developed PE.Conclusions: The combination of maternal history with low serum PAPP-A levels and abnormal uterine artery Doppler at 11-14 weeks can be used as predictor of pre-eclampsia.

Maternal outcome in obstetric ICU and HDU: a study from a teaching hospital in South India

Background: The management of critically ill obstetric patients presents a unique challenge. Dedicated High Dependency Unit (HDU) and Intensive Care Unit (ICU) for obstetric patients are widely available in India. The data regarding obstetric critical care is invaluable in formulating policy decisions. The objective is to study the profile of cases admitted to obstetric HDU and ICU and to evaluate maternal outcome and co-morbid conditions.Methods: This was a prospective observational study between January 2017 and June 2018.Results: Total number of obstetric admissions was 7966. Total admissions to obstetric ICU were 60. ICU cases accounted for 0.7% of all obstetric admissions and 1.1% all deliveries. Obstetric cases formed 1.6% of total ICU admissions. Number of admissions to HDU was 576. HDU cases accounted for 7.2 % of all obstetric admissions. HDU utilization rate was 11.32%. Hypertensive disorders of pregnancy (n=22, 33.3%), obstetric haemorrhage (n=18, 30%), septic abortion (n=2, 3.3%) were the most common conditions necessitating admission.Conclusions: Hemorrhage was the most common indication for admission to HDU. Delayed identification and referral were the important obstacles. There is a need for early booking at peripheral centres. Introduction of obstetric ICU and multidisciplinary approach has brought down the incidence of maternal mortality in present centre.

Drug utilization study of antihypertensive drugs and prevalence of blood pressure control in adult hypertensive patients based on JNC VIII guidelines in a tertiary care hospital: a cross sectional study

Background: Hypertension is a major independent risk factor for coronary artery disease, congestive heart failure, stroke, chronic kidney disease and peripheral vascular diseases if left untreated. Drug utilization study of antihypertensive drugs and the study on prevalence of blood pressure control would help in reducing the burden of the disease and health expenditure.Methods: The study was conducted in the Outpatient Department of Medicine in Government Medical College, Thrissur. Patients aged 18yrs or above diagnosed with hypertension, on antihypertensive drugs were enrolled in the study. Patients suffering from secondary hypertension and acutely ill were excluded. Patients were enrolled after taking an informed consent. Demographic data, present treatment for hypertension, associated co- morbid conditions if any, and treatment of the same were recorded. BP was recorded, and cost of treatment was calculated using CIMS.Results: A total of 250 patients were included in the study. Mono therapy was used in 64.8% patients and combination therapy in 35.2%. Overall drug utilization pattern showed that CCBs (42.8%) were most commonly prescribed, followed by ACEIs (32.4%) and ARBs (29.2%). Most commonly prescribed combination therapy was ACE I + BB (29.3%), followed by ARB + CCB (21.3%). Mean cost of antihypertensive drug therapy was 3057.8 Rs / yr. Recommended target BP was achieved in 49.6% of patients.Conclusions: The prescription pattern of antihypertensive drug was in accordance to the JNC-VIII guidelines. The blood pressure target was achieved only in less than 50% of patients.

Angiographic Aspects of Myocardial Bridges.

Introduction: The segment of epicardial coronary artery that traverses intramurally through the myocardium and bridged by a bunch of cardiac muscle fibers is called tunneled artery or intramural artery. The band of cardiac muscle fibers passing over the tunneled artery segment is named as myocardial bridge. During angiography milking effect is observed during systole due to the external pressure of muscle fibers on the tunneled artery that leads to narrowing of vessel lumen and further ischemia. Materials & Methods: It is a prospective study performed from 2012- 2015 in cardiac centers available around Tirupati, Andhra Pradesh, South India. A total number of 2015 adult patients who underwent diagnostic coronary angiography were evaluated to detect myocardial bridges. With the informed consent the relevant data was collected from the patients and analyzed. Results: The prevalence of myocardial bridges was 3.17%. Among the 2015 patients 70.7% are males and 29.2% are females. Among 64 myocardial bridge positive cases 62.5% were male and 37.5% were female patients. Regarding coronary dominance 84% were right dominant and 14.4% were left dominant and 1.6% are balanced. The percentage incidence of myocardial bridging according to dominance was 3.01% for right dominant patients, 4.12% for left dominant patients and 3.1% for balanced dominant patients. In all the myocardial bridge positive cases they were located on the left anterior descending artery (LAD). According to diagnosis the patients with normal coronaries were 22.6%, patients with MILD CAD were 17.9%, patients with single vessel disease were 23.4%, patients with two vessel disease were 14.7% and the patients with triple vessel disease were 21.3%. The 64 myocardial bridging cases were grouped in to three groups according to their age. Incidence of double bridges was observed in 3 cases of which 66.7% males & 33.3% in females. Conclusion: These results shows that Andhra Pradesh population are with high angiographic incidence of myocardial bridges (MB’s), when compared with other population in India. We observed more lengthy bridges which may cause luminal reduction of coronary vessel and myocardial ischemia (MI), we also observed higher incidence of MB’s in male patients but systolic luminal reduction is more in female patients then in males. These observations suggest that the risk of MI will be more for the female patients with MB’s.

Percentage Incidence, Morphology and Morphometry of Myocardial Bridges: A Cadaveric Study.

Introduction: Myocardial bridging is the term used when a segment of major epicardial coronary artery runs intramuscularly under the tunnel formed by fibers of myocardium that bridges instead of it’s normal or routine epicardial path. In the literature there are varying reports on clinical implications of myocardial bridges from protection against atherosclerosis to myocardial ischemia, as well as leading to infarction and sudden cardiac death. Materials and Methods: 150 adult formalin fixed human hearts which were available in the department of Anatomy and Forensic Medicine, S.V.Medical College, Tirupati, Andhra Pradesh, India. These hearts were dissected and observed for the presence, location, type, number and direction of myocardial bridges and their association with coronary dominance. With the help of digital calipers morphometric parameters (length, width& thickness) of myocardial bridges and length of blood vessel underneath the myocardial bridge were measured, noted and photographs were taken. Results: The overall incidence of myocardial bridges was 20.6%( 31/150). Among these 18.6% (28/31) were on left anterior descending (LAD) artery and 2% (03/31) were on posterior interventricular (PIV) artery. The direction of muscle fibers in the bridges were oblique to the direction of the coronary vessels in majority of cases. Length, width and thickness of myocardial bridges were in the range of 12-69.7mm, 3.74-8.6mm and 1.3-3.87mm respectively. Conclusions: Myocardial bridges may be associated with wide range of clinical problems. Contraction of myocardial bridge may results in vascular compression and myocardial ischemia. Knowledge on morphology and morphometric details of myocardial bridges facilitates cardiologists in diagnosis, planning therapeutic strategies and prognostic predictions.

Association of Calcitonin Receptor 1340 C>T Polymorphism with Periodontitis Susceptibility.

Background: Periodontitis is a common multifactorial oral disease and a major cause of tooth loss among adults. The present study was aimed to investigate the role of calcitonin receptor (CTR) gene polymorphism in the causation of periodontitis. Materials and Methods: A total of 112 subjects comprising of 62 patients and 50 controls were enrolled and recruited from various dental clinics in and around Hyderabad, India. Two milliliter of blood sample was collected from all the subjects. Following extraction of DNA, genotyping for CTR 1340 C>T was performed by PCR-RFLP. Results: The frequency of CC, CT and TT genotypes in patients was 45%, 42% and 13% while in controls it was 56%, 32% and 12%. The frequency of C and T allele was 0.66 and 0.34 in patients whereas it was 0.72 and 0.28 in controls. The genotype and allele frequencies did not vary between the groups. The genotype frequencies among male and female sub-types revealed a statistically significant difference in female subgroup. The CT genotype and T allele revealed an OR value of 5.62 and 2.40 respectively. Conclusion: Our study revealed a significant association of this SNP with periodontitis only in females. It also highlights the predisposing role of CT genotype and T allele in the causation of periodontitis. However, replicative studies on the influence of this polymorphism in different ethnic groups may identify the potentiality of this SNP towards periodontitis.

Advances in Transgenic Maize for Quality Improvement.

Maize (Zea mays) is a major food and animal feed worldwide and occupies a relevant place in the world economy and trade as an industrial grain crop. Currently more than 70% of maize production is used for food and feed; therefore, knowledge of genes involved in grain structure and chemical is important for improving the nutritional and food-making properties of maize. It is a good source of carbohydrates, fats, proteins, vitamins and minerals but deficient in two essential amino acids, Viz., lysine and tryptophan. To overcome this problem and to improve the above quality characters the maize breeders have followed different strategies like opaque 2, QPM and development of transgenic maize with improved quality characters. Finally we can conclude that the conventional breeding techniques and now plant biotechnology are helping meet the growing demand for food production, nutrition security while preserving our environment for future generations.

A study on the prevalence of refractive errors among school childern of 7-15 years age group in the field practice areas of a medical college in bangalore.

Background: Refractive Error is an avoidable cause of visual impairment. Diagnosis and Treatment of refractive errors is the simplest and most effective forms of eye care. Aims & Objective: To study the prevalence and determinants of uncorrected refractive errors, among school children of 7-15 years. Material and Methods: The study was a cross sectional study of 1378 government school children of 7- 15 years age group in both rural and urban field practice areas of Dr. BR Ambedkar Medical College, Bangalore. Students were screened for defective vision with the help of Snellen’s chart and refractionist confirmed the findings. Students with refractive error were provided with spectacles free of cost. Data was analysed to determine the prevalence of refractive errors among the school children. Results: 687 children of urban and 691 children of rural area were examined. 53.6% of the study population were boys and 46.4% were girls. The mean age of the study group was 12.4 years. The prevalence of uncorrected refractive error in urban and rural children was 7.03%. The prevalence of Myopia, Hypermetropia and Astigmatism in children was 4.4%, 1.03%, 1.6% respectively. Children 13 to years 15 attending urban schools were most likely to have uncorrected myopia. Hypermetropia was associated with younger age group and female children. Conclusion: The prevalence of uncorrected refractive error, especially myopia, was higher in older children. Causes of higher prevalence and barriers to refractive error correction services should be identified and addressed. Eye screening of school children is recommended.

Mosaic triple X syndrome in a female with primary amenorrhea.

Background: Turner's syndrome is the most common chromosomal abnormality in females, affecting 1 in 2,500 live female births. It is a result of absence of an X chromosome or the presence of a structurally abnormal X chromosome. Its most consistent clinical features are short stature and ovarian failure. Aim: The aim of the study was to report a rare case of mosaic triple X syndrome in a female with primary amenorrhea. Materials and Methods: The chromosomal analysis using GTG banding was carried out, which revealed a mosaicism with 45,XO/47,XXX chromosomal constitution. Fluorescent in situ hybridization was also carried out to further confirm the observation made in the study. Conclusion: The physical features presented by the female could be due to the 45,XO/47,XXX mosaicism and the karyotype analysis was consistent with the diagnosis and clinical symptoms. Triple X mosaicism was confirmed with conventional and molecular cytogenetic analysis.

Clinical and behavioural profile of a rare variant of klinefelter syndrome-48, XXXY.

Klinefelter’s syndrome is a sex chromosomal aneuploidy caused by an addition of X chromosome in males (47,XXY).Variants of this syndrome with X and Y polygamy are of rare occurrence. Here we describe a rare case of 48, XXXY Klinefelter’s variant from South India with a reported incidence of 1 per 17,000 to 1 per 50,000 male births. The presence of an extra X chromosome/s in these individuals has a great impact on the physical and cognitive functions, which could be attributed to gene dosage effects and genes involved in neurogenic development.

Detection of L1 (CAM) mutations in X-linked mental retardation: A study from Andhra Pradesh, India.

BACKGROUND : Mental health is an essential ingredient in the quality of life. Recent studies carried out in countries like Germany, USA, France, England and Belgium have provided evidence for the involvement of L1 (CAM) mutations in various X-linked mental retardation syndromes. L1 CAM is a neural cell adhesion molecule belonging to the superfamily of the immunoglobulins and is critical for proper CNS development in humans. AIM: This study was aimed to screen idiopathic mental retardation cases for L1 CAM mutations. MATERIALS AND METHODS : In this study, we screened 15 cases with mental retardation. Genomic DNA from the patients and control subjects was analyzed by polymerase chain reaction using specific primers. RESULTS : In 2 out of 15 patients, mutation was detected between exon 26 and 27. CONCLUSION : It is worthwhile to screen idiopathic mental retardation cases for L1 CAM mutations to reduce genetic morbidity in the population by offering genetic counseling and prenatal diagnosis.

Cytogenetic studies of 1001 Down syndrome cases from Andhra Pradesh, India.

Cytogenetic data obtained from investigating 1001 patients of Down syndrome (DS) and their parents over a period of 20 years (January 1979-January 1999) are presented. The frequency of pure trisomy, mosaicism and translocation was 87.92, 7.69 and 4.39 per cent respectively. The origin of the extra chromosome 21 due to meiotic non-disjunction was 79.24 per cent maternal and 20.76 per cent paternal. A high frequency of acrocentric chromosome associations was also observed in mothers of children of Down syndrome, this might have predisposed to an enhanced risk for non-disjunction. Birth order of DS showed a higher number of first and second borns. Reproductive performances of the parents indicated a high rate of abortions, compared to controls. Cytogenetic investigations carried out over these years greatly helped in the management of these children and for counseling the affected families.

Serum and erythrocyte magnesium levels in hypertensives and their first degree relatives.

Essential hypertension is an arbitrarily defined disorder to which both genetic and environmental factors contribute. Magnesium and its interactions with other cations may play an important role in the pathogenesis of essential hypertension. Various studies have been carried out on the levels of serum and erythrocyte magnesium in hypertensives and the results are controversial and there is no systematic study in Indian population. In the present study serum and erythrocyte magnesium levels in 86 hypertensives and their 77 first degree relatives as well as in sex and age matched controls were studied. Serum and erythrocyte magnesium levels showed a significant decrease both in the hypertensives and their first degree relatives (p < 0.01). The significantly decreased levels of magnesium in the first degree relatives suggest genetic basis of essential hypertension and may be used as marker to identify those at risk.

Erythrocyte sodium and Na+, K(+)-ATPase activity in untreated hypertensives and their first degree relatives.

In this paper we report the erythrocyte sodium concentration and Na+, K(+)-ATPase activity in 86 untreated hypertensives and their 77 first degree relatives and also in sex and age matched controls. There was significant increase in erythrocyte sodium both in the hypertensives and their first degree relatives (p < 0.01), whereas Na+, K(+)-ATPase activity was significantly reduced in the study group when compared with controls. The possibility of using these parameters as genetic markers is suggested.

Maternal reproductive history and the occurrence of Down's syndrome.

Reproductive history of mothers of 115 Down's syndrome children was studied and compared with 200 control mothers who gave birth to normal children. The frequency of spontaneous abortions in mothers of Down's syndrome babies was found to be elevated significantly (p < 0.05). The data suggest that the maternal health and reproductive potential have a prominent aetiological significance in the occurrence of Down's syndrome.

Effect of D-400, a herbomineral preparation on lipid profile, glycated haemoglobin and glucose tolerance in streptozotocin induced diabetes in rats.

This study was undertaken to investigate D-400, a herbomineral formulation in streptozotocin induced diabetes in rats. Glycated haemoglobin, lipid profile and glucose tolerance test were studied. D-400 has an established hypoglycaemic effect in alloxan induced diabetes in rats as well as in non-insulin dependent diabetes mellitus patients. D-400 treated group showed lower glycated haemoglobin, triglycerides and higher HDL levels. The hyperglycaemic response was blunted after administration of oral glucose in the same group.