Association of air pollution and homocysteine with global DNA methylation: A population-based study from North India.

BACKGROUND: Anthropogenic air pollution has been implicated in aberrant changes of DNA methylation and homocysteine increase (>15µM/L). Folate (<3 ng/mL) and vitamin B12 (<220 pg/mL) deficiencies also reduce global DNA methylation via homocysteine increase. Although B-vitamin supplements can attenuate epigenetic effects of air pollution but such understanding in population-specific studies are lacking. Hence, the present study aims to understand the role of air pollution, homocysteine, and nutritional deficiencies on methylation. METHODS: We examined cross-sectionally, homocysteine, folate, vitamin B12 (chemiluminescence) and global DNA methylation (colorimetric ELISA Assay) among 274 and 270 individuals from low- and high- polluted areas, respectively, from a single Mendelian population. Global DNA methylation results were obtained on 254 and 258 samples from low- and high- polluted areas, respectively. RESULTS: Significant decline in median global DNA methylation was seen as a result of air pollution [high-0.84 (0.37-1.97) vs. low-0.96 (0.45-2.75), p = 0.01]. High homocysteine in combination with air pollution significantly reduced global DNA methylation [high-0.71 (0.34-1.90) vs. low-0.93 (0.45-3.00), p = 0.003]. Folate deficient individuals in high polluted areas [high-0.70 (0.37-1.29) vs. low-1.21 (0.45-3.65)] showed significantly reduced global methylation levels (p = 0.007). In low polluted areas, despite folate deficiency, if normal vitamin B12 levels were maintained, global DNA methylation levels improved significantly [2.03 (0.60-5.24), p = 0.007]. Conversely, in high polluted areas despite vitamin B12 deficiency, if normal folate status was maintained, global DNA methylation status improved significantly [0.91 (0.36-1.63)] compared to vitamin B12 normal individuals [0.54 (0.26-1.13), p = 0.04]. CONCLUSIONS: High homocysteine may aggravate the effects of air pollution on DNA methylation. Vitamin B12 in low-polluted and folate in high-polluted areas may be strong determinants for changes in DNA methylation levels. The effect of air pollution on methylation levels may be reduced through inclusion of dietary or supplemented B-vitamins. This may serve as public level approach in natural settings to prevent metabolic adversities at community level.

Why parents agree or disagree for minimally invasive tissue sampling (MITS) to identify causes of death in under-five children and stillbirth in North India: a qualitative study.

BACKGROUND: Information on exact causes of death and stillbirth are limited in low and middle income countries. Minimally invasive tissue sampling (MITS) is increasingly practiced in place of autopsy across several settings. A formative research documented the experiences of counselling and consenting for MITS in north India. METHODS: This exploratory qualitative study was conducted at a tertiary care hospital in Delhi. During the early implementation of MITS, observations of the counselling and consenting process (n = 13) for under-five child death and stillbirths were conducted. In-depth interviews with MITS team members (n = 3) were also conducted. Observation and interview data were transcribed and inductively analysed using thematic content analysis to identify emerging themes and codes. RESULTS: The MITS team participated in daily ward rounds for familiarisation with parents/families. Following death declaration the counselling was done in counselling corner of the ward or adjacent corridor. Mostly the counselling was targeted at the father and family members present, using verbal explanation and the consent document in local language. The key concerns raised by parents/family were possible disfigurement, time needed and possible benefits. Most of the parents consulted family members before consent. Among those who consented, desire for next pregnancy, previous pregnancy or neonatal loss and participation of treating senior doctor were the key factors. The negative experience of hospital care, poor comprehension and distance from residence were the factors for consent refusal. Lesser number of parents of deceased children consented for MITS compared to the neonates and stillbirths. CONCLUSIONS: The initial experiences of obtaining consent for MITS were encouraging. Consent for MITS may be improved with active involvement of the treating doctors and nurses, better bereavement support, private counselling area along with improvement in quality of care and communication during hospitalisation. Special efforts and refinement in counselling are needed to improve consent for MITS in older children.

Information about exact cause of death and stillbirth are essential for appropriate care of children and pregnant women. Autopsy assists in establishing exact cause of death, but not preferred by the parents/families. Minimally invasive tissue sampling (MITS) is a suitable alternate to autopsy for establishing causes of death and stillbirth. A pilot project on MITS was initiated at a tertiary care hospital in north India (New Delhi). An exploratory formative research was conducted to document the experiences of counselling and consenting for MITS. Under this study, observation of the counselling and consenting process for conduct of MITS in under-five children (including neonates) and stillbirths were done. Additionally, in-depth interviews with MITS team members were also conducted. Counselling for MITS was done in one corner of the ward or adjacent corridor and mostly targeted at the father and family members present. Counselling was primarily verbal explanation in local language. The key concerns from parents/family were disfigurement, time needed and possible benefits. Most of the parents consulted family members before decision. Benefit during next pregnancy, past previous pregnancy or neonatal loss and senior treating doctor's involvement were key factors for consent. Poor quality of care and comprehension were the reasons for refusal.

Perceptions of family, community and religious leaders and acceptability for minimal invasive tissue sampling to identify the cause of death in under-five deaths and stillbirths in North India: a qualitative study.

BACKGROUND: Minimal invasive tissue sampling (MITS) has emerged as a suitable alternative to complete diagnostic autopsy (CDA) for determination of the cause of death (CoD), due to feasibility and acceptability issues. A formative research was conducted to document the perceptions of parents, community and religious leaders on acceptability of MITS. METHODS: This qualitative study was conducted at and around the Safdarjung Hospital, Delhi, India. Participants for in-depth interview included the parents who had either child or neonatal death or stillbirth and the key community and religious representatives. The focus group discussions (FGDs) involved community members. Process of obtaining consent for MITS was observed. Data were analyzed inductively manually for emerging themes and codes. RESULTS: A total of 104 interviews (parents of deceased children, neonates or stillbirths, n = 93; community members, n = 8 and religious leaders, n = 7), 8 FGDs (n = 72) were conducted and process of obtaining consent for MITS (n = 27) was observed. The participants were positive and expressed willingness to accept MITS. The key determinants for acceptance of MITS were: (1) understanding and willingness to know the cause of death or stillbirth, (2) experience of the healthcare received and trust, (3) the religious and sociocultural norms. Parents and community favored for MITS over CDA when needed, especially where in cases with past stillbirths and child deaths. The experience of treatment, attitude and communication from healthcare providers emerged as important for consent. The decision making process was collective involving the elders and family. No religious leader was against the procedure, as both, the respect for the deceased and need for medical care were satisfied. CONCLUSIONS: Largely, MITS appeared to be acceptable for identifying the causes of child deaths and stillbirths, if the parents and family are counseled appropriately considering the sociocultural and religious aspects. They perceived the quality of care, attitude and communication by the healthcare providers as critical factors for acceptance of MITS.

India tops the chart of childhood deaths and stillbirths globally. The causes of many childhood deaths and stillbirths remain unknown or ill-defined in India. For planning the health policies, program and clinical practices, the cause of death (CoD) and stillbirth are primarily determined by verbal autopsy (VA) method, which has several inherent limitations. The acceptance and implementation of complete diagnostic autopsy (CDA) or full autopsy for CoD determination is limited. Minimally invasive tissue sampling (MITS) is a suitable alternative to full autopsy for CoD determination. MITS has been found to be acceptable and being implemented in some low and middle income countries. Considering the multi-religion and sociocultural contexts of India, an understanding about the perceptions, practices, acceptance and barriers among the parents, community members and religious leaders regarding MITS was necessary for designing appropriate communication strategies and materials for consent. A qualitative study was undertaken in New Delhi, India using in-depth interviews with the parents of deceased children and stillbirths, community leaders, community health workers and religious leaders and focus group discussions with the community members and observation of obtaining consent for MITS. The parents and community members were positive about MITS and expressed willingness to accept it over full autopsy to know the CoD and stillbirths. The factors like past stillbirths and child deaths, experience of the healthcare received and trust, the religious and sociocultural norms emerged as important determinants for acceptance of MITS. The religious leaders were also positive about MITS if needed for treatment and benefit of the family.

Perceptions of the parents of deceased children and of healthcare providers about end-of-life communication and breaking bad news at a tertiary care public hospital in India: A qualitative exploratory study.

BACKGROUND: Parents of dying children face unique challenge and expect compassionate support from health care providers (HCPs). This study explored the experiences of the parents and HCPs about the end-of-life care and breaking bad news and related positive and negative factors in Indian context. METHODS: This qualitative exploratory study was conducted at paediatrics department of a tertiary care hospital in Delhi. In-depth interviews with the parents (n = 49) and family members (n = 21) of the children died at the hospital and HCPs (6 doctors, 6 nurses and 4 support staffs) were conducted. Also events and communication around death of eight children were observed. Data were inductively analysed using thematic content analysis method to identify emerging themes and codes. RESULTS: Doctors were the lead communicators. Majority of parents perceived the attitude, communication and language used as by resident doctors as brief, insensitive and sometimes inappropriate or negative. They perceived that the attitude and communication by senior doctor's as empathetic, positive and complete. Parents recalled the death declaration by resident doctors as non-empathetic, blunt and cold. Most parents received no emotional support from HCPs during and after death of their child. All doctors expressed that death of their patients affected them and their emotions, which they coped through different activities. The overcrowded wards, high workload, infrastructural limitation and no formal communication training added to the emotional stress of the HCPs. CONCLUSIONS: Majority of the communication by the HCPs during the hospitalisation and end-of-life period were perceived as suboptimal by the parents. The HCPs were emotionally affected and faced end-of-life communication challenges. The study highlights the communication by HCPs and support for parents during the end-of-life communication and breaking bad news. It suggests adoption of context specific communication protocol and materials and training of HCPs in communication to improve the quality of care.

Methylenetetrahydrofolate reductase gene polymorphism, global DNA methylation and blood pressure: a population based study from North India.

BACKGROUND: Hypertension is a complex disorder affected by gene-environment interactions. Methylenetetrahydrofolate reductase (MTHFR) gene is one of the genes in One Carbon Metabolic (OCM) pathway that affects both blood pressure and epigenetic phenomenon. MTHFR C677T gene polymorphism leads to reduced methylation capacity via increased homocysteine concentrations. Global DNA methylation (5mC%) also gets affected in conditions such as hypertension. However, no study is found to understand hypertension in terms of both genetics and epigenetics. The present study aims to understand the relation between methylation, MTHFR C677T gene polymorphism and hypertension. It also tries to understand relation (if any) between methylation and anti-hypertensive drugs. METHODS: This is a cross-sectional study where data were collected from a total of 1634 individuals of either sex in age group 35-65 years. Hypertensives (SBP ≥ 140 mm Hg and DBP ≥ 90 mm Hg) (on treatment/not on treatment) and absolute controls were 236 (cases) and 307 (controls), respectively. All the samples were subjected to MTHFR C677T gene polymorphism screening (PCR-RFLP) and global DNA methylation assay (ELISA based colorimetric assay). Results of both the analyses were obtained on 218 cases, 263 controls. RESULTS: Median 5mC% was relatively lower among cases (p > 0.05) compared to controls, despite controlling for confounders (age, sex, smoking, alcohol, diet) (r2-0.92, p-0.08). Cases not on medication had significantly reduced 5mC% compared to controls (p < 0.05), despite adjusting for confounders (r2-0.857, p-0.01). Among cases (irrespective of treatment), there was a significant variation in 5mC% across the three genotypes i.e. CC, CT and TT, with no such variation among controls. Cases (not on medication) with TT genotype had significantly lower methylation levels compared to the TT genotype controls and cases (on medication) (p < 0.01). CONCLUSION: Global DNA hypomethylation seems to be associated with hypertension and antihypertensive drugs seem to improve methylation. Hypertensive individuals with TT genotype but not on medication are more likely to be prone to global DNA hypomethylation. Important precursors in OCM pathway include micronutrients such as vitamin B-12, B-9 and B-6; their nutritional interventions (either dietary or supplement) may serve as strategies to prevent hypertension at population level. However, more epidemiological-longitudinal studies are needed for further validation.

Grief reaction and psychosocial impacts of child death and stillbirth on bereaved North Indian parents: A qualitative study.

BACKGROUND: Grief following stillbirth and child death are one of the most traumatic experience for parents with psychosomatic, social and economic impacts. The grief profile, severity and its impacts in Indian context are not well documented. This study documented the grief and coping experiences of the Indian parents following stillbirth and child death. METHODS: This exploratory qualitative study in Delhi (India) included in-depth interviews with parents (50 mothers and 49 fathers), who had stillbirth or child death, their family members (n = 41) and community representatives (n = 12). Eight focus group discussions were done with community members (n = 72). Inductive data analysis included thematic content analysis. Perinatal Grief Scale was used to document the mother's grief severity after 6-9 months of loss. RESULTS: The four themes emerged were grief anticipation and expression, impact of the bereavement, coping mechanism, and sociocultural norms and practices. The parents suffered from disbelief, severe pain and helplessness. Mothers expressed severe grief openly and some fainted. Fathers also had severe grief, but didn't express openly. Some parents shared self-guilt and blamed the hospital/healthcare providers, themselves or family. Majority had no/positive change in couple relationship, but few faced marital disharmony. Majority experienced sleep, eating and psychological disturbances for several weeks. Mothers coped through engaging in household work, caring other child(ren) and spiritual activities. Fathers coped through avoiding discussion and work and professional engagement. Fathers resumed work after 5-20 days and mothers took 2-6 weeks to resume household chores. Unanticipated loss, limited family support and financial strain affected the severity and duration of grief. 57.5% of all mothers and 80% mothers with stillbirth had severe grief after 6-9 months. CONCLUSIONS: Stillbirth and child death have lasting psychosomatic, social and economic impacts on parents, which are usually ignored. Sociocultural and religion appropriate bereavement support for the parents are needed to reduce the impacts.

Perceptions of the healthcare providers regarding acceptability and conduct of minimal invasive tissue sampling (MITS) to identify the cause of death in under-five deaths and stillbirths in North India: a qualitative study.

BACKGROUND: India contributes the highest share of under-five and neonatal deaths and stillbirths globally. Diagnostic autopsy, although useful for cause of death identification, have limited acceptance. Minimally invasive tissue sampling (MITS) is an alternative to autopsy for identification of the cause of death (CoD). A formative research linked to pilot MITS implementation was conducted to document the perceptions and attitudes of the healthcare professionals and the barriers for implementation. METHODS: This exploratory qualitative study conducted at a tertiary care hospital in New Delhi, India included the hospital staffs. In-depth interviews were conducted with the doctors, nurses and support staffs from pediatrics, neonatology, obstetrics and forensic medicine departments. Inductive data analysis was done to identify the emerging themes and codes. RESULTS: A total of 26 interviews (doctors, n = 10; nurses, n = 9 and support staffs, n = 7) were conducted. Almost all professional and support staffs were positive about the MITS and its advantage for CoD identification including co-existing and underlying illnesses. Some opined conduct of MITS for the cases without clear diagnosis. All participants perceived that MITS would be acceptable for parents due to the non-disfigurement and preferred by those who had unexplained child deaths or stillbirths in past. The key factors for MITS acceptance were appropriate communication, trust building, involvement of senior doctors, and engagement of the counselor prior to deaths and training of the personnel. For implementation and sustenance of MITS, involvement of the institute authority and government stakeholders would be essential. CONCLUSIONS: MITS was acceptable for the doctors, nurses and support staffs and critical for better identification of the causes of death and stillbirths. The key facilitating factors and challenges for implementing MITS at the hospital in Indian context were identified. It emphasized on appropriate skill building, counseling, system organization and buy-in from institution and health authorities for sustenance of MITS.

Metabolic distress in lipid & one carbon metabolic pathway through low vitamin B-12: a population based study from North India.

BACKGROUND: Dyslipidemia and hyper-homocysteinemia are the major independent risk factors of cardio vascular disease. Deficiency of folate and vitamin B-12 are associated with both hyper-homocysteinemia and dyslipidemia. The aim of the study is to evaluate the relationship of homocysteine and its associated dietary determinant levels (Folate and Vitamin B-12) with lipids and obesity parameters (WC, BMI, WHR) in North Indian population. METHODS: The participants were recruited under a major government funded project through household survey covering 15 villages of Haryana, India. Participants were both males and females, between age group 30-65 years, from a north Indian community. Initially 1634 individuals were recruited, of which 1374 were considered for analysis as they were not found to be on any kind of medication for high blood pressure, CAD, diabetes or any other disorder, and had no missing data. 5 mL of intravenous blood sample was collected after obtaining written informed consent from the participants. Homocysteine, folate and vitamin B12 levels were estimated through Immulite 1000 by chemi-luminescence technique. Triglyceride, total cholesterol and HDL-C were estimated by spectrophotometry technique using commercial kits. The values for LDL and VLDL were calculated using Friedwald's equation. Height, weight, waist circumference (WC), hip circumference (HC) was measured over light clothing. Statistical analysis for data was performed using SPSS 16.0 version. RESULTS: All the lipid indices, except HDL, showed a trend of negative correlation with homocysteine after controlling for confounders, though not significant. No association was found between obesity (WC, BMI, WHR) and homocysteine in the present study. Vitamin B-12 deficiency was significantly associated with both hyper-homocysteinemia and low HDL. Folate was found to have significantly reduced risk for high TC & LDL. CONCLUSIONS: The "hcy-lipid" hypothesis does not seem to be complementing in the present studied population. The population is vulnerable to severe under-nutrition due to the association of vitamin B-12 with HDL, leading to metabolic disturbance in both the pathways; lipid and one carbon metabolic pathway. Co-factors such as ethnicity, cultural practices, and lifestyle & dietary habits must be considered while making public health policies to control diseases.

Association of DD genotype of angiotensin-converting enzyme gene (I/D) polymorphism with hypertension among a North Indian population.

Hypertension, a major risk factor for cardiovascular diseases, is among the leading causes of morbidity and mortality worldwide. Genetic predisposition to the risk of developing hypertension due to angiotensin-converting enzyme (ACE) gene insertion(I)/deletion(D) polymorphism (through altered serum ACE activity) is well documented among various populations. The present study investigated the possible association between ACE (DD) genotype and hypertension using a nested case-control study design including 451 individuals (of either sex in the age group 30-65 years) from a rural North Indian population practicing agriculture and lacto-vegetarianism. Blood Pressure was classified using JNC-7 criterion. Age- and sex-matched individuals were selected from normotensive (N-122), pre-hypertensive (N-123), hypertensive not on medication (N-122), and hypertensive on medication (N-84) categories. Amplification of DNA and genotyping of PCR product was done using standard protocols. From the analysis, comparatively higher frequency of individuals with DD genotype in the hypertensive category was observed, indicating a possible relation between DD genotype and hypertension. The odds ratio analysis revealed 2.225 (1.13-4.37)-fold significant increased risk for hypertension among cases, validating the vulnerability of individuals with DD genotype towards hypertension. Thus, the present study highlights the increased risk for developing hypertension due to ACE DD genotype in the studied population.