Saumya Pandey

Immunobiology of Toll-like receptors: Emerging trends

Toll‐like receptors (TLR), a family of evolutionarily conserved pathogen recognition receptors, play pivotal role as primary sensors of invading pathogens. TLR identify molecular motifs of infectious agents (pathogen associated molecular patterns) and elicit an effective defensive response against them. Mammalian TLR derive their name from the Drosophila Toll protein, with which they share sequence similarity. So far, expression of 10 TLR is known in humans. The adaptor proteins, myeloid differentiation factor 88 and Toll IL‐1 receptor (TIR) domain containing adaptor inducing IFN‐β (TRIF) are the key players in the TLR signalling cascade leading to the activation of nuclear factor (NF)‐κB and interferon regulatory factor‐3, respectively. Targeted manipulation of the TLR signalling pathway has immense therapeutic potential and may eventually prove to be a boon in the development of innovative treatments for diverse disease conditions. There is accumulating evidence that TLR agonists have tremendous potential as novel therapeutic targets. In this review, we have discussed the immunobiology of TLR and emphasize significant advances made within the ever‐expanding field of TLR that provide intriguing insights efficacious in unravelling the complexities associated with TLR.

Targeting Wnt-Frizzled signaling in cardiovascular diseases

Wnts are secreted glycoproteins implicated in biological processes ranging from embryonic cardiac development to uncontrolled cell proliferation in diseased conditions. Cardiovascular disease is a major cause of morbidity and mortality worldwide. Phenotypic modulation of vascular smooth muscle cells, migration and proliferation in intimal layer and increased extracellular matrix production are some of the known hallmarks of cardiovascular pathologies. Heterogeneity associated with the binding of Wnts to their transmembrane receptors, Frizzled, and coreceptors low density lipoprotein-receptor-related protein is indeed intriguing. Nuclear-cytoplasmic shuttling of beta-catenin and activation of transcriptional factors, lymphoid enhancer factor and T cell activation factor leading to target gene activation has remained elusive. Our review highlights the emerging role of Wnt-Frizzled signaling in cardiovascular diseases. Overall, the pathway appears to be an attractive therapeutic target in identifying susceptible individuals at risk of developing restenosis/other vascular pathologies in the near future.

Re: Effect of locally‐tailored labour management guidelines on intrahospital stillbirths and birth asphyxia at the referral hospital of Zanzibar: a quasi‐experimental pre–post‐study (The PartoMa study)

pain has been resolved. We agree with the authors that the cause of a haemoperitoneum in pregnancy (and its accompanying abdominal pain) is sometimes hard to distinguish pre-operatively, because of the nonspecific clinical presentation and numerous differential diagnoses that can be considered; however, imaging modalities like ultrasound sonography or magnetic resonance imaging can be helpful to quantify the extent of the intraabdominal haemorrhage, and in specific cases the origin of the bleeding. The actual management of a haemoperitoneum greatly depends upon the clinical presentation, the findings at physical examination, the results from laboratory or imaging investigations, and the gestational age. Moreover, the identification of risk factors, like the use of controlled ovarian hyperstimulation for artificial reproductive techniques before pregnancy and a history of (deep) endometriosis, can be helpful in the recognition of SHiP and may affect the choice of treatment. We agree that a surgical approach is often unavoidable, but expectant management (combined with fluid resuscitation) can be considered when signs of hypovolemic shock or fetal distress are absent. We agree that more research is needed to gain further insight into this rare phenomenon, and therefore an international registration network is currently recording cases of SHiP prospectively (International Network of Obstetric Survey Systems, INOSS). By a detailed analysis of these cases more insight into the pathological mechanism of SHiP, and the required interventions needed to improve fetomaternal and perinatal outcomes, will hopefully be obtained.&

Re: Caesarean scar pregnancy in the UK: a national cohort study

Sir, We would like to add that the elegantly conceptualised article by Harb et al., assessing the clinical outcomes and stringent management of caesarean scar pregnancies (CSP) in a well-defined patient population subset in the UK, is informative. Pregnancyrelated complications and clinical management protocols are indeed essential to cost-effectively reduce the morbidity and mortality associated with reproductive disorders, including miscarriages, CSP and infertility. The study involved 86 participating early pregnancy units with confirmed CSP diagnosis between November 2013 and January 2015 using a monthly mailing system as part of the UK Early Pregnancy Surveillance Service (UKEPSS). An accurate estimate of 102 cases of CSP was accomplished, with an estimated incidence of 1.5 per 10 000 (95% CI 1.1–1.9) maternities. The meticulously designed clinical research study convincingly demonstrated that the success rates of expectant, medical and surgical management were 43, 46 and 96%, respectively; interestingly, the complication rates were 71% with expectant, 60% with medical and 36% with surgical management, whereas discharge from care (median number of days) was 82% (range 37–174) with expectant, 21% (range 10–31) with medical, and 11% (range 4–49) with surgical management. The overwhelming data suggest that surgical management appears to be associated with a high success rate, low complication rate, and short post-treatment follow up; we wish to comment that the study data sets may be further evaluated in stratified/subgroup cases with relevant statistical adjustments for covariates, including dietary patterns, tobacco usage and partner’s/husband’s smoking status so as to have better correlations of clinical outcomes of CSP. Further, a case–control gene epidemiological study with age-matched nonpregnant women and pregnant women with caesarean scar pregnancy may be conducted so as to eventually validate biomarkers for CSP management in women of ethnically disparate populations, including the UK cohort. Future research is warranted to provide cost-effective strategies for dissecting the underlying cellular and molecular mechanisms associated with CLP in ethnically disparate patient populations.&

Re: Immediate versus delayed initiation of the levonorgestrel-releasing intrauterine system following medical termination of pregnancy - 1-year continuation rates: a randomised controlled trial: Levonorgestrel-releasing intrauterine system following medical termination of pregnancy: a randomised controlled trial.

Sir, We thank Dr Saumya Pandey for her positive and interesting remarks. The purpose of our paper was to contribute to the scarce knowledge regarding women’s use of antenatal healthcare services and interventions during childbirth in pregnancies subsequent to stillbirth. Hopefully, such knowledge provides a base for future studies aiming to evaluate the effects of care and interventions that currently are or should be provided for this patient group. Our study took place in a highincome country were antenatal care is free of charge for all women. We certainly agree that accessible and high quality maternal healthcare is essential in reducing maternal and fetal mortality and morbidity, particularly from a global perspective. However, what constitutes cost-effective healthcare in pregnancies following stillbirth remains to be defined. In terms of reducing the risk of recurrent fetal demise and other adverse pregnancy complications, the evidence base regarding optimal management of subsequent pregnancies is limited. Furthermore, although additional antenatal appointments and technological surveillance are often welcomed by parents, the effectiveness of increased monitoring in reducing psychological distress is not proven. Additional care specifically addressing the psychological needs of parents seems to be less often provided.&

Re: Healthcare utilisation, induced labour and caesarean section in the pregnancy after stillbirth: a prospective study

Sir, The recently published article by Gravensteen et al. investigating healthcare utilisation, induced labour, and caesarean section (CS) in the Norwegian Mother–Child Cohort Study (901 pregnant women) is indeed interesting and provides meaningful clinical insights in women’s health research. Stillbirth is a major public health problem worldwide; direct causes of maternal deaths and stillbirth are primarily due to obstetric complications, primarily haemorrhage, sepsis, miscarriages, complications of recurrent spontaneous abortions, preeclampsia and eclampsia, and prolonged/obstructed labour. I would like to comment that the authors have conceptualized, designed, and successfully executed this population-based pregnancy cohort study by incorporating a large sample size (174 pregnant after stillbirth, 362 pregnant after live birth, and 365 previously nulliparous); a relatively larger sample size (901 pregnant women) definitely adds to the study strengths by increasing the statistical significance and power of the study, thereby emphasizing the accuracy of data derived from questionnaires in the second/third trimesters of pregnancy and clinical history extracted from the Medical Birth Registry of Norway. Moreover, informed consent is a core tenet of good practice research, and the authors have adhered to the guidelines of bioethics during the enrolment of eligible subjects for research. The stringent inclusion criteria for study subjects of a specific population subset (Norwegian ethnicity) further enhances the overall study quality by ruling out any potential bias and/or possibility of population admixture while critically evaluating the main study outcomes viz. self-reported assessment of antenatal care and register-based assessment of onset and mode of delivery. The findings of this elegant study involving reproductive-aged women suggest that women with a previous stillbirth had a comparatively higher number of antenatal visits (mean 10.0; 95% CI 9.4– 10.7) than women with a previous live birth (6.0; 5.8–6.2) and previously nulliparous women (6.3; 6.1–6.6). Interestingly, subgroup stratification with relevant adjustment of covariates/statistical parameters revealed that the prevalence of induced labour and CS, elective and emergency, was significantly higher in the stillbirth group. Adequate, patient-friendly, one-to-one counselling sessions informing pregnant women about the risk factors associated with pregnancy-related complications are warranted to effectively address the growing phenomenon of psychosocial distress and grief associated with stillbirth among pregnant women at antenatal visits. In the present study, the authors demonstrate that anxiety was a minor mediator for association between stillbirth and frequency of antenatal visits; on the contrary, dread of childbirth was not a significant mediator for elective CS. Furthermore, Norwegian women pregnant after stillbirth make more use of healthcare services and more often had induced labour and CS. To conclude, I strongly feel that costeffective maternal healthcare service utilization is critical for identifying community-level determinants and psychosocial factors in the management of pregnant women of diverse ethnicities so as to significantly reduce the morbidity and mortality associated with pregnancy-related complications, including stillbirth. Effective strategic and collaborative monitoring, both preand post-pregnancy, is urgently required to massively improve maternal and fetal outcomes by providing good health care services on a global platform.&

Re: Genetic variation in the progesterone receptor gene and susceptibility to recurrent pregnancy loss: a case–control study

Sir, The recently published article by Bahia et al. assessing the association of progesterone receptor (PGR) gene variants with susceptibility to recurrent pregnancy loss (RPL) provides critical clinical research insights in reproductive medicine. Pregnancy-related complications, including stillbirths, miscarriages and infertility, are emerging as major public health challenges among ethnically disparate populations worldwide, including women of Indian ethnicity. We wish to comment that the authors have elegantly conceptualised this pregnancy-related cohort study by incorporating a retrospective case–control gene epidemiological design with eligible participants enrolled from outpatient obstetrics/gynaecology clinics; the stringent inclusion/exclusion criteria added to study merits, and the core tenets of ethical research involving informed consent of participants were followed during the entire course of the study. A relatively higher sample size of cases (women with RPL 396) and controls (women 361) increased statistical power; moreover, RPL was defined as three or more consecutive miscarriages of unknown aetiology among pregnant women. An allelic exclusion method (real-time PCR) for PGR genotyping yielded high-quality, reliable results assessing PGR single nucleotide polymorphisms and comparative allelic, genotypic and haplotypic distributions. A major study strength was the sophisticated, selection-bias free, robust statistical data analyses using linkage disequilibrium, minor allele frequency and homozygous versus heterogyzous genotypic frequency distribution(s) in cases versus controls by multiple comparisons and adequate adjustments of co-variates/parameters, namely age, body mass index and menarche; as active clinical medicine researchers in reproductive medicine, we would like to suggest that the authors could have included further clinically relevant covariates for data analysis such as tobacco usage (chewers versus smokers), vitamin D deficiency, caffeine and alcohol usage, Mycobacterium tuberculosis and human papillomavirus positivity among RPL cases so as to draw more definitive conclusions, and accordingly devise cost-effective, predictive biomarkers in RPL management among women of reproductive age from varying genetic landscapes and sociocultural as well as socio-economic backgrounds. The findings demonstrated higher minor allele frequency of rs590688, rs10895068 and rs1942836 in women with RPL than in control women; interestingly, association analysis further indicated significantly higher frequencies of heterozygous (1/2) rs608995, along with heterozygous (1/2) and homozygous (2/ 2) rs590688, rs10895068 and rs1942836 genotype carriers between women with RPL versus control women, respectively. An increased risk of RPL associated with rs590688 and rs1942836 was dependent on the number of minor alleles, thereby implicating a ‘dose-dependent’ effect associated with both genetic variants; linkage disequilibrium values were accurate and rs590688, rs10895068, rs608995 and rs1942836 PGR variants were significantly associated with RPL. Overall, the study strongly implicated positive association of specific PGR variants (rs590688, rs10895068 and rs1942836), and PGR haplotypes (ATGCCGTC, ATTCGGTC) with increased risk of RPL, thereby emphasising the emerging role of PGR as an RPL candidate locus. Future cohort-based studies amalgamating immunodiagnostics, electrophysiology, radioimaging modalities and cellbased assays coupled with transcriptomics, genomics, metabolomics and proteomics would prove to be immensely beneficial in the identification/validation of predictive biomarkers in identifying RPL-susceptible women of diverse ethnicities. A multicentric, collaborative clinical management protocol is warranted to provide a robust public health research model in strategically addressing pregnancy-related metabolic disorders and/or complications in both the developing nations as well as lowresource countries.&

Re: Partner smoking influences whether mothers quit smoking during pregnancy: a prospective cohort study

Sir, The recently published article by Rom an G alvez et al. assessing the prevalence and intensity of cigarette smoking among pregnant women and their partners, and modifiable factors associated with quitting smoking provides critical clinical research insights inobstetrics-gynaecology. Tobacco consumption (smoking or chewing) is a significant predictor of metabolic perturbations in reproductive physiology, including pregnancy-related complications/stillbirths/miscarriages/infertility, and is emerging as a major public health problem in ethnically disparate populations worldwide. We would like to comment that the authors have elegantly conceptualized this tobacco-related prospective cohort study in pregnant women (n = 486) of Andalusia, Southern Spain; stringent inclusion/ exclusion criteria added to study merits, and core tenets of good practice research/ bioethics including informed consentwere adequately followed. Interestingly, the follow-up of study cohort in first/second/ third trimesters of pregnancy was welldesigned; a relatively large sample size of 486 pregnant women increased statistical power. The sound research methodology involving thorough estimation of proportions of women and partners who quit smoking at each trimester was impressive; tobacco consumption in the form of the number cigarettes per day throughout pregnancy was stringently investigated among pregnant women/their partners. As active clinical medicine researchers in the highly competitive, ever-expanding reproductive medicine/healthcare management field(s), we wish to suggest that the authors could have included few clinically relevant covariates for subgroup stratification amongmild/moderate/heavy smokers, viz. serum cotinine levels, VitaminDdeficiency, caffeine/alcohol/carbonated drink intake, bone mineral density, diet (vegetarian/eggetarian/non-vegetarian), Anti-Mullerian hormone level(s), menarche, blood group, marital years, psychiatric disorders/anxiety/depression, Mycobacterium tuberculi/ Human papillomavirus positivity among pregnant women so as to draw more definitive conclusions, anddevise cost-effective community-based tobacco cessation-pregnancy management guidelines with public health research models in specific population subsets of reproductive-aged pregnant women of varying genetic landscapes and socio-economic strata. Main outcomes involved determination of factors associated in multivariable model considering socio-demographic/obstetric/ anthropometric/lifestyle variables of pregnant women and smoking habits of partners; 61.08% of women quit smoking during pregnancy (95% CI 53.61–68.55) while smoking rate amongmothers diminshed from 36.06% (n = 167) before pregnancy to 14.08% (n = 65), 12.39% (n = 54) and 11.92% (n = 51) during three pregnancy trimesters (P < 0.001). Moreover, the intensity of tobacco consumption decreased from 8.71 cigarettes/day in first to 5.51 in second (P < 0.001) and 5.96 cigarettes/day in third trimester (P = 0.0002 first vs. third). A minimal decrease in smoking frequency among partners was observed, and women whose partner smokedwere relatively less likely to quit (adjusted odds ration (aOR) 0.26, 95% CI 0.12–0.55). The results provided meaningful insights in tobacco cessation research protocols and indicate that one in ten pregnant women smoked, and one in four was a passive smoker; the findings strongly advocated that tobacco exposure reduction strategies in pregnancy should stringently focus on partner smoking. Future multicentric tobacco control/ prevention healthcare management public health research models incorporating larger sample sizes utilizing pooled study cohorts of diverse ethnicities, including Spanish, British, Danish, North American, Latin American, African, Australian and Indian (including Indian pregnant women/their partners at Indira IVF Center, Udaipur, Rajasthan) coupled with psychosocial interventions are warranted efficiently to address health risks and/or psychosocial distress associated with cigarette smoking trends among pregnant women and their partners.&