The cross-sectional study design was conducted in collaboration with the Emergency Medicine, Clinical Psychiatry, Hospital Administration and Department of Biochemistry at a four-decade old tertiary care teaching hospital affiliated to medical school in South India. The authors began recruiting the cases of Deliberate Self-Harm (DSH) for comprehensive clinical and laboratory evaluations of various blood and biochemical parameters as soon as they were admitted in emergency or critical care unit.

The study was approved (254/Ethics Committee/KIMS/NKP/2019) by the institutional ethical committee of KIMS Medical College before beginning the data collection. The authors began the recruitment and analysis from 1^st January 2019 till 31^st December 2019 for a duration of 12 months. This helped in improving the modest sample size of 202 patients to achieve minimum sample error of no more than 5% (95% confidence). DSM-IV-TR based psychiatric diagnosis were only recruited and assessed [12].

Inclusion and Exclusion criteria: The authors included either gender, above age of 18 years and of all socio-economic status using Modified Kuppuswamy Scale [13] were evaluated for biochemical abnormalities to all those who had been attempting suicide but failed and could give their consent for the study irrespective of their medical or psychiatric diagnosis. The refusal of the consent for evaluation was respected and there was strict confidentiality maintained on data sheet.

Blood evaluation and analysis were done on the same day of hospitalisation for complete blood counts, serum electrolytes, proteins and cholesterol. To measure the blood cholesterol level, 5 mL of blood sample was collected after at least 8 hours of fasting from the subjects and analysis was done on BS-390 automated Chemistry analyser using cholesterol oxidase peroxidase method [14]. The authors used the following standardised Instruments for evaluation: Schedules for Clinical Assessments in Neuropsychiatry (SCAN) [15]; Structured Clinical Interview for DSM disorders Clinical Version-1 (SCID CV-1); Deliberate Self Harm Inventory [16]; International Personality Disorder Examination; Beck’s Suicide Intent Scale [17]; Socio-demographic and clinical profile sheet.

Data obtained were analysed using SPSS 21.0 version of software. The appropriate descriptive (frequency, percentage, mean) as well as inferential (for degree of association, Pearson’s correlation and multivariate regression analysis for identifying the covariance of suicidality with serum cholesterol and other biochemical abnormalities like serum proteins, blood pH and electrolyte disturbances) statistical tests like Chi-square test for qualitative variables were applied. For all tests of probability, p-value less than 0.05 was considered significant.

A total of 202 patients were evaluated in the present study, among them 120 (59.4%) were males while remaining 82 (40.6%) were females with 111 (54.9%) falling in the age range of 20-29 years. Further it observed as the age progressed the frequency of suicidal attempts reduced, however there was very minimal difference of 25 (12.4%) vs 22 (10.9%) prevalence between 40-49 years and more than 50 years respectively [Table/Fig-1]. This suggests two changing trends, one is suicidal attempts are getting common in employed as well as unemployed and Indian families are getting differentiated into nuclear families not only in urban and metro cities but in rural India as well. As far as the clinical scenario of mode of deliberate harm is concerned as seen in [Table/Fig-2], poisoning 178 (88.1%) was preferred mode followed by hanging 14 (6.9%) irrespective of genders. Out of the 202 patients in the study, 175 patients came in with first ever suicide attempt or deliberate harm attempters. It was observed that 22 (10.9%) had second attempt over a period of 6 months and 5 (2.5%) individuals had multiple suicidal attempts. When the variety of the intent of most of the patients was analysed 83 (41.1%) had moderate intent followed by severe intent in about 64 (31.7%) of patients.

The dispersion of psychiatric diagnosis in suicide attempters were mainly depressive disorder, substance use disorder, dual diagnostics, bipolar disorder, Borderline Personality Disorder (BPD) formed major chunk of population as shown in [Table/Fig-3]. But interestingly 13 (6.4%) patients were also having debilitating medical diseases like cancer(s) while 4 (2.0%) patients had intense anxiety (panic) attacks. Suicidal attempts were relatively more common in schizophrenia 7 (3.5%) than delusional disorders 4 (2.0%). Almost 169 (83.7%) of patients had normal complete blood counts but 106 (52.5%) showed acidic pH (<7.3) while only 5 (2.5%) of patients had alkalosis. Serum sodium levels were abnormal only in 16 (7.9%) of patients, while serum potassium was abnormal in 43 (21.3%) of patients. Further serum chloride was deranged in 13 (6.4%) while serum [Table/Fig-4] albumin was deranged in 17 (8.4%). About 13.4% (27 out of 202 patients) were found to have increased serum cholesterol while rest had surprisingly normal levels of cholesterol and almost similar number of patients had abnormal liver function test. The authors observed a strong and significant (p=<0.01) positive association with serum potassium, serum cholesterol and serum pH levels, while others were insignificant [Table/Fig-5]. In furthering inferential statistical analysis as reflected in [Table/Fig-6], the multinomial regression method generalises logistic regression to multiclass problems, for example with more than two possible discrete outcomes which occurs with suicidal patients. We know that age and gender have been found to be important predictors of suicidality and the authors chose additional three more factors which can influence the likelihood ratio of suicidal behaviour in patients of DSH like frequency of deliberate harms, severity of intent, level of education apart from the serum cholesterol levels. The likelihood covariance was highest and clearly significant with increased serum cholesterol (Chi-square value: 26.084; df: 10; p*=0.004), closely followed by suicidal intent just before the attempt (Chi-square value: 17.490; df: 10; p=0.064) and effect of age (Chi-square value: 40.588; df: 10; p=0.094).

Since there are no other studies in suicidal patients for abnormal biochemical parameters like pH, serum electrolyte, serum proteins and most of the focus had remained on levels of cholesterol, the present discussion has been related to debate on cholesterol levels in suicidal neurobiology. Excessive and dampened reactivity of the neurobiological stress system have been associated with increased vulnerability for psychiatric disorders [18]. The present study demonstrated that in contrast to the conventional understanding of low serum cholesterol diathesis a newer trend of analysis was indeed required which this study bridges. The study evaluates all possible routine biochemical blood parameters with more emphasis on serum cholesterol levels of all suicidal patients visiting emergency services.

A meta-analysis by Wu S et al., in 2016 of as many as 65 epidemiological studies using predominantly Cochrane database from 1980 to 2014 using random effect model with a total sample of 510,392 participants clearly found that suicidal patients had significantly lower total cholesterol (WAD:- 24.75, 95% CI- 27.71 to -21.78) [19]. Further they reported that lower serum total cholesterol levels were associated with 123% higher risk of suicide attempt (95% of CI: 24% to 302%). However, the major limitation of their meta-analysis was potential heterogeneity, narrow range of cholesterol depletion and there was not a single Indian study included to compare. Further, obesity prevalence has increased significantly every decade. Therefore, in the present study authors have attempted to re-examine the real effect of cholesterol on suicidality from rural tertiary care health services from south India. We have evaluated all prospective suicidal attempters visiting our tertiary health care set up in emergency medicine services for identifying if serum cholesterol affects suicidal behaviour, not only as risk factor for depressive mood state but irrespective of any psychiatric diagnosis which makes the present study clearly different from many others [20-22]. However, the present study has the conforming comparisons for blood cholesterol levels with few other studies [23-26].

Interestingly, there are studies which demonstrated that lower serum cholesterol levels are also linked with major depressive disorder (OR 4.229 CI 95% 2.555-7.000, p<0.001) and suicide attempt (OR 5.540 CI 95% 2.825-10.866, p<0.001) in their study by Segoviano-Mendoza M et al., [27]. Similarly, a study by Morgan RE et al., published in lancet that plasma cholesterol and depressive symptoms in older men found that depression was three times more common in the group with low plasma cholesterol (<4.14 mmol/L) than in those with higher concentrations (5/31 (16%) vs 22/363 (6%); p=0.033) [28]. In support of hypocholesterolemia and impulsive suicidality in BPD, a study by Atmaca M et al., in 16 BPD found that mean cholesterol and leptin levels of the patient group were significantly (p<0.05) lower than those of the controls [29]. In another study, [30] in 60 psychotic patients of which only 13 patients attempted suicide, just marginal and insignificant (p-value=0.91) change of mean serum cholesterol level was reported in their results, though they highlighted it as significant positive predictor.

Another interesting study from Poland evaluated 148 schizophrenic suicidal patient and assessed serum cholesterol along with LDL, HDL and triglycerides and reflected that in depressed patients, low cholesterol can be state- dependent risk factor for suicidal behaviour [31]. Even para-suicidal schizophrenics have also reported with lower cholesterol levels [32,33]. On the contrary, Huang T et al., 2000 in Taiwanese population of schizophrenics found no difference in serum cholesterol levels irrespective of suicidal attempt [34]. On contrary, a study from Korea found correlation between low cholesterol and suicidality holding true only for depression but not for schizophrenia [35]. And on neutral/equivocal levels another group of investigators compared serum lipid levels of 282 schizophrenics, 72 bipolar disorders and 242 major depressive disorders and concluded that biological levels of serum lipids do not predict suicide risk [36]. Finally, like the present study findings, an earlier study from India by John S et al., did not support the use of biological indicators such as serum total cholesterol HDL LDL or TAG either to predict any functional psychiatric outcome including suicidality [37].

Compared to most previous studies [6,20,22,23,27], why the present study suicidal patients have not shown lower serum cholesterol levels? One of the reasons could be that epidemiologically violent suicide attempts were found more commonly with low cholesterol levels thus in the present study most of the suicidal attempters were perhaps not having violent suicidality compared to western countries. Another reason could be huge intergroup variation based on psychopathologies and metabolic status in suicide attempters. As seen in [Table/Fig-6] of the present study, the multinomial regression reflects serum levels of cholesterol as one of the significant biochemical abnormality in suicidal patients. The support for this also comes from a recently published case control study from Italy in 2019 [38]. Over recent years, in usual clinical practice, most psychiatric patients are following sedentary lifestyles and they have less control over eating; many of them avoid physical exercise and modest number of them have weight gain, metabolic or cardiac problems.

The present study has certain limitations which needs to be considered as main issue of approach in identifying the biochemical abnormalities including serum cholesterol should never been just a mere cross-sectional or hospital-based study as method of evaluation. The practical inability to assess the fluctuation of blood cholesterol with the fluctuation of mood, impulsivity, sleep deprivation status, type of diet one had or missed just before the suicide attempt which might impair its direct causal association as noted in current findings always remains difficult domains to be controlled.

This is an encouraging study supporting notion that though dyslipidemia influences suicidal behaviour than other biochemical abnormalities, there is still the lack of convincing power of cholesterol as a putative biomarker for suicidal behaviour and more research is required in this vast field to determine the neurochemistry of suicidal behaviour. This can only be made possible by building for national healthcare database from India to have one homogeneous voice rather than discrete formulation of changing hypothesis. Its indeed a nature’s puzzle which keeps us pondering for perfect molecular diagnostic answer as there are many psychological conditions and the on-going medications for treatment of medical disorders which might influence one’s eating and exercise habits, and subsequently cholesterol levels. There is also a possibility that cholesterol might be a bystander of the association between suicide and some of these factors as we conclude that no single biological marker may be sufficiently accurate for complex suicidal behaviour prediction.