Cytokine gene polymorphisms and sudden infant death syndrome.

AIM: Several studies indicate that the mucosal immune system is stimulated in cases of sudden infant death syndrome (SIDS), and our hypothesis is that this immune reaction is because of an unfavourable combination of functional polymorphisms in the cytokine genes. METHODS: Thus, in this study, single nucleotide polymorphisms (SNPs) in the genes encoding IL-6, IL-8, IL-12, IL-13, IL-16, IL-18 and IFNgamma were investigated in 148 SIDS cases, 56 borderline SIDS cases, 41 cases of infectious death and 131 controls. RESULTS: Regarding genotype distribution, no differences between the investigated groups were found. However, in the SIDS group, the genotypes IL-8 -251AA/AT and IL-8 -781CT/TT were significantly more frequent in the SIDS cases found dead in a prone sleeping position, compared with SIDS cases found dead in other sleeping positions. In addition, there was an association between fever prior to death and the genotype IL-13 +4464GG in the cases of infectious death. CONCLUSION: This study indicates that specific interleukin genotypes are a part of a genetic make up that make infants sleeping prone at risk for SIDS.

Bacteriological investigation--significance of time lapse after death.

In cases of sudden unexpected death in infants and children (SUDI), microbiological investigation has been an important part of the autopsy protocol at the University of Oslo for the last 15 years. The purpose of this study was to compare the microbiological findings in samples taken at hospital admittance shortly after death and at autopsy. Blood cultures and cerebrospinal fluid (CSF) were collected both at the hospital and at autopsy; organ samples were additionally collected at autopsy. Hospital samples were collected at a median of 4.5 h (95% confidence interval [CI] 3.25-5) and autopsy samples at a median of 24.25 h (95% CI 22-25.5) after death. The proportion of positive cultures was stable over time; the post mortal time had no influence on bacterial growth. As long as the autopsy is performed within 48 h after death, prior microbiological examination is unnecessary. Blood culture, CSF and lung specimens are the best predictors in our study.

Determination of cotinine in pericardial fluid and whole blood by liquid chromatography-tandem mass spectrometry.

Cotinine is the main metabolite of nicotine and is used as an indicator of exposure to tobacco smoke. A method has been developed for quantification of cotinine in pericardial fluid and whole blood collected from autopsy casework involving cases of infant death. Sample clean-up was achieved by solid-phase extraction with a mixed-mode column. Cotinine was quantified by liquid chromatography-tandem mass spectrometry. Positive ionization was performed in the multiple reaction monitoring mode. Two transitions were monitored for the analyte and one for the internal standard, cotinine-d(3). The calibration range was 0.9-176 ng/mL for cotinine in both matrixes. The recovery of the analyte ranged from 86 to 92%, and the between-assay precisions ranged from 4 to 6% relative standard deviation. Whole blood and pericardial fluid samples from 95 infant deaths obtained during autopsy were analyzed. A strong correlation (R(2) = 0.97) was found between the cotinine concentrations in pericardial fluid and blood. The correlation was not affected by the postmortem time interval. This study demonstrates that pericardial fluid may be an alternative specimen to blood for quantification of cotinine in forensic autopsies.

Are substitutions in the first hypervariable region of the mitochondrial DNA displacement-loop in sudden infant death syndrome due to maternal inheritance?

AIM: To investigate whether all substitutions in the first hypervariable region (HVR1) in sudden infant death syndrome (SIDS) can be recovered along the maternal line of the family (inherited), or whether SIDS victims have new substitutions compared to maternal relatives (somatic mutations) that may be related to environmental factors. METHODS: Seventy-one SIDS/mother pairs, including 11 families with SIDS, mother and mother's relatives and/or SIDS siblings, were studied. The HVR1 sequence was recorded in the base-pair range 16056-16400. The recorded HVR1 sequence was compared with the Cambridge sequence, and differences were recorded as substitutions. The substitution pattern in the SIDS victims was compared with the pattern found in family members along the maternal line. RESULTS: All the substitutions found in SIDS victims could be traced in the maternal line of the family; in 5 cases this was observed through three generations, and in 3 cases through four generations. DISCUSSION: In patients with known mitochondrial (mt) DNA disease, a large number of sequence variants have been found in the D-loop region. Substitutions in the D-loop may be part of a haplotype with mutations elsewhere in the mtDNA. CONCLUSION: HVR1 substitutions in SIDS victims are hereditary and not due to somatic mutations.

Comparative epidemiology of sudden infant death syndrome and sudden intrauterine unexplained death.

BACKGROUND: Unexplained antepartum stillbirth and sudden infant death syndrome (SIDS) are major contributors to perinatal and infant mortality in the western world. A relation between them has been suggested. As an equivalent of SIDS, only cases validated by post mortem examination are diagnosed as sudden intrauterine unexplained death (SIUD). OBJECTIVE: To test the hypothesis that SIDS and SIUD have common risk factors. METHODS: Registration comprised all stillbirths in Oslo and all infant deaths in Oslo and the neighbouring county, Akershus, Norway during 1986-1995. Seventy six cases of SIUD and 78 of SIDS were found, along with 582 random controls surviving infancy, all singletons. Odds ratios were obtained by multiple logistic regression analysis. RESULTS: Whereas SIUD was associated with high maternal age, overweight/obesity, smoking, and low education, SIDS was associated with low maternal age, smoking, male sex, multiparity, proteinuria during pregnancy, and fundal height exceeding +2 SD. Thus the effects of maternal age were opposite in SIUD and SIDS (adjusted odds ratio 1.39 (95% confidence interval 1.17 to 1.66) per year, p < 0.0005). Heavy smoking, male sex, and a multiparous mother was less likely in SIUD than in SIDS (0.22 (0.06 to 0.83), 0.22 (0.07 to 0.78), and 0.03 (<0.01 to 0.17) respectively). Overweight/obesity and low fundal height were more common in SIUD than in SIDS (7.45 (1.49 to 37.3) and 13.8 (1.56 to 122) respectively). CONCLUSIONS: The differences in risk factors do not support the hypothesis that SIDS and SIUD have similar determinants in maternal or fetal characteristics detectable by basic antenatal care.

[Finding the cause of death in intrauterine death--which examination should be done?]. / Påvisning av dødsårsak ved intrauterin død--hvilke undersøkelser bør gjøres?

BACKGROUND: The incidence of antepartum fetal death after 22 weeks of gestation is about 0.4% in Oslo, Norway. Screening routines will decide how many cases will remain unexplained. One quarter are diagnosed as Sudden Intrauterine Unexplained Death (SIUD). Precise knowledge of the cause of death is needed as a basis for counselling, prevention and treatment. MATERIAL AND METHODS: The implementation of diagnostic routines in stillbirths in Oslo from 1986 to 1995 was examined. A structured review has been prepared and new guidelines for diagnostic procedures have been adopted by The Perinatal Committees of Oslo and Akershus. RESULTS: Autopsies and placental investigations were performed in 88% of intrauterine deaths. Among SIUD cases (all autopsied), infectious causes were examined and excluded in 93%. In contrast Kleihauer-Betke test was performed in only 17%, autoantibodies explored in 24%, and glucose tolerance tested in 36% of cases. The recommended laboratory investigations are: autopsy, placental investigation, bacteriological culture and investigations of infections such as Toxoplasma gondii, cytomegalovirus, parvovirus B19 and Listeria monocytogenes, Kleihauer-Betke test, screening for diabetes, chromosome analysis of amniotic fluid and placental biopsy, and serology of antiphospholipids. INTERPRETATION: Both the content and implementation of diagnostic routines in the cases of antepartum fetal death has not been optimal in our region; there is need for a change.

Mitochondrial DNA point mutations detected in four cases of sudden infant death syndrome.

The aim of this study was to investigate the tRNA(Leu(UUR)) gene and the first part of the ND1 gene in mitochondrial DNA (mtDNA) in cases of sudden infant death syndrome (SIDS). A total of 158 cases of SIDS and 97 controls were included in the study, and the base pairs in the range 3230-3330 were investigated using polymerase chain reaction (PCR) and temporal temperature gradient electrophoresis (TTGE). If a band shift was detected by TTGE, the area investigated and the D-loop was sequenced. Three different point mutations (T3290C, T3308C and T3308G) were detected in four of the SIDS cases, while none of the controls were mutated. We also found a high D-loop substitution rate in these four cases. The findings indicate that mtDNA mutations may play a role in some cases of SIDS.

Inflammatory responses in sudden infant death syndrome -- past and present views.

Sudden infant death syndrome (SIDS) is sudden unexpected death in infancy for which there is no explanation based on commonly accepted diagnostic criteria; however, half of the victims have had slight signs of infection prior to death. Such slight infection with fever is an important risk factor in combination with a prone sleeping position, especially in infants between 2 and 4 months of age. The purpose of this review is to summarise findings that support the theory that a significant part of cot deaths may be due to an overreaction to otherwise harmless infections. Such factors are mucosal immune stimulation, cytokines in the cerebrospinal fluid and hypoxanthine levels in vitreous humour. The review aims at explaining why we believe that a slight infection combined with a prone position, a warm environment and a vulnerable age period may trigger a vicious circle leading to death.

IL-6 cerebrospinal fluid levels are related to laryngeal IgA and epithelial HLA-DR response in sudden infant death syndrome.

The objective was to investigate whether there is any correlation between signs of central and peripheral immune stimulation in victims of sudden infant death syndrome (SIDS), the former expressed by IL-6 in cerebrospinal fluid (CSF), the latter by IgA, IgG, and IgM immunocytes, T lymphocytes, and HLA-DR expression in laryngeal mucosa. Seventeen SIDS cases with low CSF IL-6 levels (< or =5 pg/mL) and 20 cases with high CSF IL-6 levels (> or =30 pg/mL) were subjected to immunohistochemical quantitation of IgA, IgG, and IgM immunocytes; semiquantitative scoring of T lymphocytes in the mucosa of epiglottis and larynx, and semiquantitative evaluation of HLA-DR expression. SIDS cases with IL-6 levels > or =30 pg/mL had a significantly higher number of IgA immunocytes in laryngeal mucosa (p = 0.007) and in epiglottis (p = 0.03) than cases with IL-6 levels < or =5 pg/mL. Furthermore, laryngeal HLA-DR expression was significantly more extensive in SIDS cases with IL-6 levels > or =30 pg/mL than in those with levels < or =5 pg/mL (p = 0.05). No differences were found for IgG and IgM immunocytes or for T cells. In addition, babies found prone more often had symptoms of slight infection before death and had a higher number of IgA immunocytes in the larynx (p = 0.02) than babies sleeping on their side or back. Because IL-6 levels > or =30 pg/mL correspond to the levels found in infants who die from infectious diseases such as meningitis/septicemia and pneumonia, the findings favor the hypothesis that many SIDS cases may be caused by an "overreaction" of the immune system to an otherwise harmless infection.

The complement component C4 in sudden infant death.

UNLABELLED: The aim of the present study was to compare partial deletions of the complement C4 gene in victims of totally unexplained sudden infant death (SID) (n = 89) and borderline SID (n = 15) with and without slight infections prior to death, in cases of infectious death (n = 19), and in living infants with and without infections (n = 84). The SID and borderline SID groups were pooled. In this total SID group slight infections prior to death was associated with deletion of either the C4A or the C4B gene (P = 0.033), and the SID victims with such infections had a higher deletion frequency than the controls (P = 0.039). There were no differences between the living infants with and without upper airway infections. CONCLUSION: The present study confirms that partial deletions of the C4 gene in combination with slight upper airway infections may be a risk factor in sudden infant death.

Increased number of substitutions in the D-loop of mitochondrial DNA in the sudden infant death syndrome.

The purpose of the present study was to investigate substitutions in the D-loop of mitochondrial DNA (mtDNA) in sudden infant death syndrome (SIDS) and controls, since several observations indicate the involvement of mtDNA mutations in SIDS. These include elevated levels of vitreous humour hypoxanthine in SIDS victims, familial clustering without mendelian traits, and observations of increased sleepiness and a lower activity score in infants who later succumbed to SIDS. Eighty-two cases of SIDS and 133 controls were investigated and the D-loop sequences were recorded in the base-pair range 16055-16500 in the mtDNA sequence. The sequencing was carried out using the Applied Biosystems Sequenase dye terminator method and a ABD373A sequencer. The recorded D-loop sequences were compared with the Cambridge sequence and differences were recorded as substitutions. The SIDS cases had a tendency towards a higher substitution rate in the D-loop than the controls (p = 0.088). This observation makes it interesting to search for deleterious mutations in other locations in the mtDNA.

Are elevated cerebrospinal fluid levels of IL-6 in sudden unexplained deaths, infectious deaths and deaths due to heart/lung disease in infants and children due to hypoxia?

Many SIDS cases probably die after periods of hypoxia and it has been shown that hypoxia may stimulate IL-6 production. The purpose of this paper was to examine if there were any correlations between hypoxanthine in vitreous humour and IL-6 in CSF. The concentration of IL-6, IL-1beta and TNFalpha in cerebrospinal fluid of 50 Sudden Infant Death syndrome (SIDS) cases, 9 borderline SIDS cases, 18 infectious deaths, 8 violent deaths and 22 cases with heart/lung disease were measured by ELISA. The hypoxanthine (Hx) vitreous humour concentrations in the same groups were determined by high performance liquid chromatography. The IL-6 levels in cases of infectious death, heart/lung disease and borderline cases were significantly higher than in the SIDS cases (p < 0.01). The Hx levels were in the same range in cases of SIDS, borderline SIDS and infectious death, and they were significantly higher than the levels in cases of violent death and heart/lung disease (p < 0.01). There was no correlation between hypoxanthine and IL-6 in any of the groups. In the cases studied IL-6 elevation is probably not induced by hypoxia, but is rather a result of immunological stimulation.

Hypoxanthine levels in vitreous humor: a study of influencing factors in sudden infant death syndrome.

Elevated hypoxanthine (Hx) levels in the vitreous humor of victims of the sudden infant death syndrome (SIDS) have been been claimed to indicate hypoxia before death. An important methodologic problem is the temperature-dependent Hx increase after death. Although most previous studies have corrected for expected postmortem Hx increase, the aim of the present study was to omit this problem by matching the subjects of the different groups studied for similar postmortem time. Thirteen cases of violent death, 11 cases of borderline SIDS, 22 cases of death from infectious disease, and 17 cases of death from heart/lung disease were compared with a 3-fold number of SIDS cases. To investigate the impact of environmental factors on the Hx levels of the SIDS cases, the following possible confounding factors were investigated: attempt to resuscitate, slight infections before death, sleeping position, and time of the day when found dead. To investigate whether the size of the eyeball would influence the Hx level, five cases of violent death in children were compared with 15 adults who died suddenly. The vitreous humor Hx level was significantly higher in SIDS victims than in cases of violent death (p < 0.01) and deaths due to heart/lung disease (p < 0.01), whereas no such difference was found either between SIDS and borderline SIDS or SIDS and infectious death. There were no differences between any of the subgroups of SIDS victims. The vitreous humor Hx level in small children and adults did not differ. The findings raise the question of possible similarities in death mechanism in SIDS and infectious death.

Objective measurements of nicotine exposure in victims of sudden infant death syndrome and in other unexpected child deaths.

OBJECTIVE: Self-reported maternal smoking is associated with a dose-related-increase in the risk of sudden infant death syndrome (SIDS). The aim of this study was to measure objectively whether victims of SIDS are more exposed to tobacco smoke before death than infants who die unexpectedly of other causes. DESIGN: Continine levels in pericardial fluid were used as an indicator of exposure. Levels > 5 ng/mL indicated significant exposure, and levels > 20 ng/mL indicated heavy exposure. Samples were obtained from all sudden deaths in children < 7 years of age that occurred from 1990 through 1993 in southeastern Norway. Twenty four infants died of SIDS, 12 infants of infections, and 9 of accidents (median age 4.5, 5, and 35 months, respectively). RESULTS: Compared with the age-matched infectious deaths, a significantly higher proportion of victims of SIDS had been significantly (92% vs 67%) or heavily exposed (25% vs 0%) to nicotine, (P < .05). Median cotinine levels in infants with SIDS, 15.8 ng/mL, were significantly higher than in infants who had infectious deaths 7.1 ng/mL (P < .003) but were comparable to those of accident victims (12.9 ng/mL, not significant). CONCLUSIONS: Victims of SIDS are more often and more heavily exposed to tobacco smoke doses before death than are infants who have sudden infectious deaths. Accidental death in infancy and childhood is often associated with a significant exposure to nicotine.

SIDS--changes in the epidemiological pattern in eastern Norway 1984-1996.

The objective was to analyse differences in the epidemiological pattern of sudden death in infancy during two time periods--the Sudden Infant Death Syndrome (SIDS) 'epidemic': 1984-1989, and the period of rapid decline in the SIDS rate 1990-1996. Sex distribution, age, sleeping position, signs of infection, day of the week and place of death were registered and compared for the two time periods studied in all SIDS cases autopsied at the Institute of Forensic Medicine, Oslo. There were significantly more deaths in the age group under four months in the period 1984-89 than in the second period. Prone sleeping position, signs of infection, death outdoors and during the winter were more frequent during the first period than in the second. These features also were more frequent in the age group under four months than in the older babies during the first period. The shift in the epidemiological pattern after 1990, when the risk factor campaign was launched, indicates that prone sleeping position, cold climate, sleeping outdoors and infections seem to be risk factors that are particularly harmful to the youngest infants.

[Forensic autopsy after possible medical malpractice. A 3-year material from the Institute of Forensic Medicine in Oslo, 1993-95]. / Rettsmedisinsk obduksjon ved spørsmål om medisinsk feilbehandling. Et treårsmateriale fra Rettsmedisinsk institutt i Oslo, 1993-95.

According to Norwegian law, cases where medical diagnostic procedures or treatment may have caused death shall be reported to the police. During the period 1993-1995, an autopsy was performed at the Institute of Forensic Medicine, Oslo, in 76 cases of such deaths in hospital. In only one case did the police investigation result in a sentence for malpractice. The chief county medical officers react more often; in 16% of the cases, hospital procedures were criticised, or the hospitals were advised to improve hospital routines, and another 14% of the cases were reported to the Norwegian Board of Health. Thus, as expected, the chief county medical officers react more often with criticism than the police do with accusation. The appearance of the police in the hospital often implies a serious conflict of cultures. There are good arguments for specialists in forensic medicine acting as intermediaries between the health service and the judicial system.

Use of new Nordic criteria for classification of SIDS to re-evaluate diagnoses of sudden unexpected infant death in the Nordic countries.

To investigate whether changes in diagnostic practice might be the cause of the SIDS epidemic in the Nordic countries in the 1970s and 1980s a cooperative study was initiated in 1990. Common morphologic diagnostic criteria for SIDS were established in 1992 and 127 randomly selected sudden unexpected infant deaths from all Nordic countries from 1970 to 1995 and 205 cases from the Institute of Forensic Medicine, Oslo, Norway (RMI) from 1984 to 1995 were re-evaluated blindly using the new criteria. Neither the increase nor the decline in the SIDS rate since 1989 seemed to be due to changed diagnostic practices. SIDS seemed to have been under-diagnosed before the new criteria came into operation in 1992. There were fewer discrepancies between the original and revised diagnoses in the RMI cases than in the rest of the Norwegian cases, both before and after 1992.

SIDS cases have increased levels of interleukin-6 in cerebrospinal fluid.

Cerebrospinal fluid (CSF) from 20 infants who died of sudden infant death syndrome (SIDS), 7 cases of infectious death and 5 cases of violent death were examined with respect to concentrations of interleukin-6 (IL-6). The measurements were performed by ELISA. IL-6 levels in SIDS were significantly lower than in infectious death (p < 0.02), but significantly higher than in violent death (p < 0.02). Since IL-6 plays an important role in immune responses and may induce fever, the findings may suggest that immune activation plays a role in SIDS. The presence of cytokines in the central nervous system (CNS) may cause respiratory depression, especially in vulnerable infants.