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DOI: http://dx.doi.org/10.20534/ESR-16-9.10-145-148
Shodiev Gafur Baratovich, Assistant
of theTashkent Medical Academy,Uzbekistan Tukhtaev Kadir Rakhimovich, MD, PhD, professor of the Tashkent Medical Academy, Uzbekistan E-mail: [email protected] Giyasov Zayniddin Asomovich, MD, PhD, professor of the Tashkent Medical Academy, Uzbekistan
Diagnostic value of the lung surfactant in sudden infant death syndrome (SIDS)
Abstract: Morphological, physic-chemical and immunohistochemical properties of the lung surfactant from 50 corpses of infants aged 1 month to 1 year were studied. In the babies with sudden infant death syndrome (SIDS) the physic-chemical studies have shown pronounced decrease of the surfactant stability. Immunohistochemistry revealed decreased expression of surfactant protein B in most cases of SIDS, which is probably due to qualitative changes in lung surfactant. The comprehensive studies of lung surfactant of infants may contribute to clarifying the degree of risk and more accurately determining the causes of death in some cases of sudden infant death. This will help reduce the risk of diagnostic errors, i.e., hypo- or over-diagnosis SIDS and is of practical importance for forensic examination.
Keywords: Sudden infant death syndrome (SIDS); lung surfactant; surfactant stability; surfactant protein B; immuno-histochemistry.
Introduction
Sudden infant death syndrome (SIDS) includes sudden, nonviolent deaths of infants aged 1 month to 1 year. In this syndrome results of scene investigation, review of the clinical history, autopsy, morphological and other laboratory studies do not provide clear evidence about the cause of infant death. This issue is of great interest not only for pediatricians but for pathologists, forensic scientists and other professionals. To date, there are many hypotheses, dedicated to the aetiology and pathogenesis of sudden infant death syndrome. However, based on a critical analysis of all existing hypotheses, P.N. Goldwater (2011) rightly points out that there is not such a hypothesis, which would meet all the requirements of pediatricians, pathologists, lawyers and etc. [6]. He believes that every hypothesis must consider pathological and epidemiological risk factors. Many authors agree that SIDS is a complex syndrome that may have multiple etiologies with a lot number of possible risk factors. In their view, great prospects in this area are new approaches to identifying predisposing to SIDS risk factors disclosure molecular genetic mechanisms of this syndrome [1; 3; 6; 8; 13].
Despite the ongoing debate, one of the most common is respiratory hypothesis that infant death due to sudden occurrence of apnea. It is believed that sleep apnea may be the primary genesis, due to lack of regulatory function of the respiratory center of the brain stem, or it may be secondary to reflex the influence of a diseased lung feedback type [15]. Unfortunately, the background to the emergence of apnea and pulmonary possible mechanisms to ensure conditions for a sudden apnea, remain unknown. Almost all the researchers are unanimous in the fact that when detected in the lungs of SIDS moderate morphological changes is not enough to cause reflex respiratory arrest [5; 11; 12; 13;14]. Promising in this respect are the studies of lung surfactant system (LSS), which plays a leading role in the respiratory function of the organism. Quantitative and functional changes of LSS may entail profound disturbances in the respiratory system, including death. LSS condition at SIDS has been
studied by some authors [4; 9; 10]. Unfortunately these works were made using different, not comparable research methods and their results are contradictory. Only some immunohistochemical studies have appeared, indicating a significant decrease in surfactant protein type A (SP-A) in children with SIDS in the first months after birth [16]. Nevertheless, the role of the LSS in the pathogenesis of SIDS remains substantially open. We hypothesized that the cause of death in some cases of SIDS could actually be a sudden development of secondary apnea caused by unknown until structural and functional changes in the lungs. We assume that these changes may be primarily associated with the hidden congenital or acquired deficiency of lung surfactant in the whole or its individual components (proteins of type B, C and A, phospholipids and others). All this leads to the urgency and the need for further comprehensive research status of LSS in children with SIDS using morphological, physic-chemical and immunohistochemical methods. The data obtained will undoubtedly contribute to the optimization of the differential diagnosis of lung diseases in children, identifying high-risk groups to the development of SIDS with further development ofpreventive measures, and, last but not least, clarification of the place and importance of LSS in a highly complex pathogenesis of this syndrome.
The aim of this work was a comprehensive study of the morphological, physic-chemical and immunohistochemical features of the lung surfactant system (LSS) in SIDS, comparing them with other cases of sudden infant death with a verified diagnosis.
Material and Methods
The material for the study was collected from 50 corpses of children aged 1 month to 1 year. Of these, in 20 cases there was SIDS (I group). 18 children patients died of pneumonia and acute respiratory infection (II group). In 12 cases was a category ofviolent death (mechanical asphyxia, mechanical injuries and thermal burns - III group). The last two (II and III) groups served as controls for SIDS. For morphological studies of lung tissue taken within 2-12 hours after death were fixed in 12% formalin solution in phosphate
buffer (pH - 7,3) and after the respective wires embedded in paraffin. Paraffin sections 5-7 microns thick were stained with hematoxylin-eosin and Schiff's reagent. To identify of pneumocytes type II (P2) sections were stained by Sudan III. Counting the number of P2 in each case carried out by 1000 all pneumocytes and expressed in absolute numbers and percentages.
The surface-active properties of the lung surfactant were examined in broncho-alveolar lavage (BAL) by modified methods Langmuir-Wilhelm [9]. The minimum and maximum surface tension (ST . and ST ) of surfactant film expressed in milliNewtons
v min max' L
/ m (mN / M), from which was calculated the surfactant stability index (SI).
Immunohistochemical detection of mature surfactant protein B-type was performed on paraffin sections of lungs using a set UltraVision (Thermo Scientific, USA). These studies were conducted in 10 samples of lung SIDS, 5 - in cases ofpneumonia and acute respiratory infections, and 5 - in the categories ofviolent death. Evaluation of the results of immunohistochemical studies were conducted on a scale ofintensity and localization ofthe reaction product. All digital data is processed by the method ofvariation statistics using the software package Microsoft Excel 2010. Differences satisfying P <0.05 were considered significant.
Results
Macroscopic examination of the lung tissue with SIDS in some cases revealed the presence of foci of punctate hemorrhages under the visceral pleura, segmental pulmonary edema with emphysema-
tous areas. A microscopic examination revealed a slight swelling and minor bleeding lesions of lung parenchyma, and pulmonary vascular engorgement in violation of the permeability of the walls and diapedesis of red blood cells in the surrounding tissue. In the lumen of the large bronchi determined locally homogeneous mass of reddish color, and in some cases showed a small peribronchial infiltrates consisting mainly of lymphocytes. Small pockets of advanced alveolar atelectasis and disatelectasis have been found frequently. Often in the alveoli and bronchi increased epithelial desquamation, and sometimes the presence in the lumen of the alveoli small hemorrhagic exudates were revealed. Approximately one-third of infant deaths from SIDS in the respiratory passages and the alveoli found signs of moderate catarrh. Along with them in about one-third of deaths in SIDS morphological manifestations of lung immaturity have been found. Microscopic changes in the lungs of the control groups of infants (II and III) are fully consistent with the diagnosis and cause of death, revealed by the expert investigation.
Thus, in some cases of SIDS in the lung tissue are revealed slight morphological changes with characteristics of moderate serous-des-quamative or serous-catarrhal pneumonia. These changes are not enough to qualify as a major cause of death. However, it is possible that they may play a role as provoking factor (trigger) that causes sudden respiratory failure during sleep with outcome to death.
The results of quantification indexes of type II pneumocytes (P2) in the lung tissue are shown in Table 1.
Table 1. - Quantitative indicators of type II pneumocytes (P2)
The studied groups n The absolute number of type II pneumocytes % of type II pneumocytes
Sudden Infant Death Syndrome,SIDS (I group) 20 78±12,5* 7,8
Pneumonia and acute respiratory viral infection, ARVI (II group) 18 92,7±6,8 9,2
Cases of violent death (III group) 12 115,5±6,2 11,5
Note: the sign * marked significant changes as compared with the IIIgroups (P <0.05).
Quantitative analysis of P2 showed that babies who died a violent death (III group), they range from 9 to 12% of all cells. Absolute number in this group ranged from 87 to 122 per 1000 pneumocytes lung tissue that was equal to an average of115.5 ± 6.2. These figures we have taken as a basic reference, since they reflect the true number of P2 healthy babies.
Several other data obtained in the group of infants who died from pneumonia and ARVI (II group). The number of P2 in this group ranged from 6 to 10%, and their absolute amount varied from 67 to 118, which was equal to 92.7 ± 6.8 per 1000 lung tissue cells on average. As can be seen from the above figures, when pneumonia and ARVI, the number of P2, though somewhat reduced compared
(P> 0.05). It can be explained by the increase in frequency of lung tissue dysfunction pneumocytes in general and in particular P2, in infants who died from pneumonia and ARVI.
The number of P2 in infants with SIDS significantly fluctuated widely, ranging from 5 to 10% of all cells. The absolute number of more than 50% in cases ranged from 48 to 75 per 1000 lung tissue cells. In other cases, the number was more than 80, and in three cases reached 110-120 per 1000 cells. The average absolute number of P2 was 78 ± 12.5 per 1000 cells, which was significantly lower than in group III with a violent death (P <0.05).
Research results biophysical indicators surface activity of lung surfactant for SIDS and control groups are shown in Table 2.
to Group III, however, this decrease was not of a significant nature
Table 2. - The biophysical indicators surface activity of lung surfactant (M±m)
The studied groups n Minimum surface tension (STJ Maximum surface tension (ST ) N max' Surfactant stability index (SSI)
Sudden Infant Death Syndrome,SIDS (I group) 20 27,3±0,85* 41,6±1,3 0,41±0,02*
Pneumonia and acute respiratory viral infection, ARVI (II group 18 17,8±0,92 39,5±1,1 0,76±0,08
Cases of violent death (III group) 12 14.6±0,93 40,2±1,6 0,93±0,04
Note: the sign * marked significant changes as compared with the II and IIIgroups (P <0.05).
As can be seen from the table 2, the children who died a violent death, lung tissue has a high surface activity of surfactant. This is evidenced by low rates of ST . and high index of surfactant
/ min o
stability(SSI). In the group of children who died of inflammatory lung diseases, it was a slight increase in STmin, and, accordingly, a slight decrease in rates of surfactant stability. However, these changes were not statistically significant. The most pronounced changes in the biophysical properties of lung surfactant discovered in SIDS. Indicator of STmin in most children of this group, appeared enlarged. Surfactant stability index (SSI) was significantly lower compared with the control groups.
Immunohistochemical studies have shown that surfactant protein is localized in the surface of the alveoli and terminal bronchioles and bronchi ofvarious calibers. Typically, the reaction product was localized in the form of aggregates, its diffuse distribution on the surface of the airways is determined very rarely. In the lung tissue of infants of the II and III groups showed rather intense and widespread deposition of surfactant protein B. At the same time, in the lung tissue in 6 of the 10 infants with SIDS expression of surfactant protein B was negative or very low compared to the control groups. This is indicative of the qualitative changes of lung surfactant, which generally confirmed the results of our biophysical researches.
Discussion
Our histological studies in approximately one-third of infant deaths from SIDS showed signs of moderate catarrhal inflammation in the respiratory passages and alveoli. Along with them, in about one-third of deaths in SIDS morphological manifestations of lung immaturity has been revealed [17]. However, these changes were not enough to qualify as a major cause of death. In respiratory function, especially in the age of 2 to 4 months, plays a special role of pulmonary surfactant. Structural and functional impairments of the pulmonary surfactant lead to aggravation of hypoxia, which is the basis of many respiratory diseases in children and adults [2]. Study of surfactant system of the lungs in SIDS devoted few works. Some researchers in the broncho-alveolar lavage (BAL) of SIDS infants found various changes in the chemical composition of the surfactant in the form of reducing the concentration of phosphatidylcholine, violation ratio of phospholipids/proteolipids, reducing the physical properties of surfactant [9; 10]. In contrast, other researchers did not find any significant changes in surfactant, which could explain the cause of sudden infant death [4].
Our studies showed that the vast majority of children with SIDS had marked impairment of the functional properties of lung surfactant. These changes are manifested in the form of disruption of the surface-active properties of lung surfactant, significant reduction in performance stability. Using immunohistochemistry method A. Stray-Pedersen, A.Vege, A.Stray-Pedersen, et al.
(2008), have found that children with SIDS in the first months observed a significant decrease in surfactant protein-A [16]. It is these periods of life coincide with the peak of the highest infant death. Based on this, the authors had been hypothesized that the reduction of SP-A is a one of risk factor for SIDS. We have also found that SIDS in some cases accompanied by severe reduced number of pneumocytes type II. It is known that pneumocyte type II (P2) is the main source of phospholipid and protein components of the lung surfactant [2]. Lack of maturity, reduced number and impaired functional activity of pneumocytes type II, of course, can lead to quantitative and qualitative changes in lung surfactant. By method of immunohistochemistry we also have found decreased expression of surfactant protein B in the majority of children with SIDS compared to infants who died of pneumonia or in accidents. However, in violation of the lung surfactant system cannot exhaustively explain the cause of death in SIDS. As we are discussed above, many authors are deciding that the prenatal or perinatal defects of regulatory mechanisms of respiratory or/and cardiovascular centres of the brain stem in SIDS have no conclusive evidences [7]. In this regard, the concepts of multifactorial causation with interaction of risk factors with variable probabilities are less restrictive and more in keeping with the large number and varying prevalence of demonstrated risk factors [8]. In our opinion, the SIDS has undoubtedly multi-etiologic character with the sophisticated multi-level pathogenesis. In the development of the syndrome very important role are playing multiple risk factors arising in the prenatal and postnatal periods in the life of infants [1]. Based on these data, we also believe that impaired lung surfactant system is just one of the risk factors, which creates a kind of unfavorable background. Against this background, under certain conditions (wrong position the baby during sleep, lack of oxygen in the room, parental smoking and other risk factors), it is possible worsening hypoxia neurons of the brain stem. This, in turn, leads to a negative reflex effect, contributing to infringement of self-repair mechanisms (autoresuscitation) with sudden death. It is not excluded that this same hypoxic depression has an adverse effect on the cardio-vascular center and causes impaired heart function up to sudden heart failure.
Conclusions
In the most children with SIDS revealed pronounced changes in the physic-chemical and immunnohistochemical properties of lung surfactant. The comprehensive studies of lung surfactant of infants may contribute to clarifying the degree of risk and more accurately determining the causes of death in some cases of sudden infant death. This will help reduce the risk of diagnostic errors, i.e., hypo- or over-diagnosis SIDS and is of practical importance for forensic examination.
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DOI: http://dx.doi.org/10.20534/ESR-16-9.10-148-150
Ergashev Nasriddin Shamsiddinovich, Head of the Department of Hospital Pediatric Surgery and Oncology TashPMI Otamuradov Furkat Abdukarimovich, Senior Research Fellow-Competitor of the Department of Hospital Pediatric Surgery and Oncology TashPMI. Ergashev Nargiza Nasriddinovna, Senior Research Fellow-Competitor of the Department of Neurology of child neurology TashPMI, 1 Tashkent Pediatric Medical Institute, Republic of Uzbekistan, Tashkent E-mail: [email protected]
Anomalies of spine and spinal cord in children with anorectal malformations
Abstract: Associated malformations with anorectal malformations (ARM) are observed in various combinations, in 1060,7% of combined anomalies of the spine and spinal cord. Objective: to determine the nature of concomitant pathology of the spine, spinal cord and related disorders in children workstation. Results: We included 51 children after surgery for various forms ofARM carried out in the clinic 2004-2015year. The prevalence of functional disorders (60.8%) in normal and satisfactory anatomical structure (76.5%) of the surveyed data prompted us to study after surgery. The results showed that neurospinal cord anomalies/dysraphism (ND) or sacrovertebral anomalies/dysraphism (SD) are widespread at the workstation, which confirms the need for a focused examination of each patient for diseases of the spine and spinal cord, the observation and treatment of co pediatric surgeon and a neurologist.
Keywords: anorectal malformations, Associated anomalies of the spine and spinal cord, diagnostics, children.
Anorectal malformations (ARM) — one of the many groups Objective: to determine the nature of concomitant pathology
proctologic pathology, requiring surgical treatment. The high mor- of the spine, spinal cord and related disorders in children undergoing
tality rate is not typical for this group of anomalies, but after surgical surgery for anorectal malformations.
correction offunctional disorders observed in 30-60% of cases. The Materials and methods. The study included 51 children
situation is compounded by the presence of associated anomalies (35 girls with, boys — 16) aged from 1 day to 15 years after sur-
that the risk ofviolations of social adaptation of patients. Associated gery for ARM performed in the clinic in 2004-2015. Preoperative
malformations observed at the workstation in a variety of combina- diagnostic measures were aimed at establishing forms of anomalies,
tions in 10-60.7% of the combined anomalies of the spine and spinal evaluation of anatomical and functional condition of the perineum
cord [1; 2; 4; 6; 7; 8]. Although the pathogenesis ofARM remains and sphincter apparatus of the rectum (contrast irgography, ultra-
completely obscure their superior combination of them with the sound of internal organs). Related osteoneural spine abnormalities,
pathology of the spine associated distal common embryogenesis [3; spinal cord and spinal hidden dysraphism installed on the results
10]. A more informative methods of diagnosis of these anomalies of the inspection and consultation neurologist, additional studies
are ultrasound and MRI of the lumbosacral spine [5; 9]. of the spine spondylography digital (Siemens) or MSCT (before
