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Gastrointestinal dysfunction and disaccharide intolerance in children infected with human immunodeficiency virus
Gastrointestinal dysfunction and disaccharide intolerance in children infected with human immunodeficiency virus R. H. Y o l k e n , MD, W. Hart, I. O u n g , MD, C. Shiff, MD, J. G r e e n s o n , MD, a n d J. A. P e r m a n , MD From the Department of Pediatrics, and Pathology, Johns Hopkins School of Medicine, the Department of Immunology and Infectious Diseases, Johns Hopkins School of Public Health, and the Department of Pediatrics, Mount Washington Hospital, Baltimore, Maryland
Because gastrointestinal dysfunction is a major problem in children with human immunodeficiency virus (HIV) infection, we utilized breath hydrogen measurements to determine the relationship between disaccharide malabsorption and gastrointestinal dysfunction in HIV-infected children. We found a strong association between lactose intolerance and persistent diarrheal disease in this population (p <0.007, Mann-Whitney U test). We also found evidence of sucrose malabsorption and persistent diarrheal disease in three of the children. Extensive microbiologic evaluations failed to reveal an etiologic agent related to the occurrence of gastrointestinal symptoms. Our findings indicate that disaccharide intolerance is a common occurrence in HIV-infected children with persistent diarrheal disease. Careful attention to dietary intake may be required to ameliorate clinical symptoms and to maintain a d e q u a t e nutrition. (J PEDIATR 1991;118:359-63)
Human immunodeficiency virus replicates in the lymphocytes and macrophages of infected persons; many of the clinical manifestations of HIV infection are due to immune cell dysfunction and subsequent immunodeficiency. 1 However, it is being increasingly recognized that HIV infection results in a multisystem disorder and that dysfunction of other organ systems can contribute to the disease state. 2 One such organ system is the gastrointestinal tract. Acute and chronic diarrhea, weight loss, abdominal pain, esophagitis, and other gastrointestinal symptoms have been noted in a substantial portion of adults with HIV infection. 35 In adults these symptoms are often, but not invariably,
Supported by grant R01-DK-40540 from the National Institutes of Health. Submitted for publication Aug. 24, 1990; accepted Oct. 1, 1990. Reprint requests: Robert H. Y01ken, MD, Pediatric Infectious Diseases, CMSC 1109, Johns Hopkins Hospital, Baltimore, MD 21205. 9/20/25781
associated with infections with gastrointestinal pathogens such as Cryptosporidium, Giardia lamblia, Mycobacteriurn avium-intracellulare, cytomegalovirus, and other agents.6, 7 Gastrointestinal dysfunction can predispose infants and young children to electrolyte imbalance and malnutrition, which may further exacerbate the T cell defects associated with HIV replication.8, 9 Furthermore, the reliHIV
Human immunodeficiency virus
[
ance of infants on milk products as the principal source of nutrition might make infants particularly susceptible to the effects of disaccharidase deficiency and other absorptive abnormalities related to epithelial cell damage. Severe gastrointestinal dysfunction and malnutrition have been reported in children with HIV infection, 1~ 11 but the mechanisms involved have not been elucidated. We utilized breath hydrogen measurements to evaluate carbohydrate absorption prospectively in HIV-infected infants and children.
359
360
Yolken et al.
METHODS Patient population. The study population consisted of 17 infants with documented HIV infection who were followed at Johns Hopkins Hospital and Mount Washington Pediatric Hospital, both located in Baltimore, Md. All children met the criteria for HIV infection in children as defined by persistently positive antibody titers for HIV for longer than 15 months (as documented by enzyme-linked immunosorbent assay and Western blot analyses), or, in the cas~of younger children, by an acquired immunodeficieney syndrome-defining illness or by the identification of HIV in peripheral blood lymphocytes. 12 Patients ranged in age from 4 to 80 months. Seven of the children were receiving 3'-azido-2',3'-dideoxythymidine at the time of analysis. Clinical data were collected from the patients by standard methods. Patients were judged to have persistent diarrheal disease if they had more than three watery bowel movements per day for a minimum of 2 weeks. Informed consent was obtained from the parents or legal guardians of all study subjects. The study protocols were approved by the Johns Hopkins School of Medicine Joint Committee on Human Experimentation. Breath hydrogen analyses. Breath H2 was measured after oral lactose administration with the use of previously described methods. 13 Patients fasted for either 6 hours, in the case of infants less than 1 year of age, or 8 hours in the case of older children. The children were fed lactose at a dosage of 2 g m / k g (maximum50 gm) in the form of a 20% aqueous lactose solution. Three of the patients were also examined for breath H2 generated after the oral administration of 20% aqueous solutions of sucrose at a dosage of 2 gm/kg. Breath sample s were collected by the nasal prong technique at 0, 30, 60, 90, 120, 150, and 180 minutes after disaccharide ingestion. 13 Breath samples were stored in 30 ml syringes for analysis on the day of collection, or in specialized 20 ml evacuated rubber-stoppered glass tubes if analysis was deferred to a subsequent day. Hydrogen concentration was determined in parts per million by gas chromatography at 25 ~ C with a Quintron detector (Quintron Instruments, Milwaukee, Wis.). Each result was corrected to an alveolar H2 concentration with carbon dioxide as an internal standard. 14 The results of each assay were expressed in terms of the peak corrected H2 concentration above the baseline value (delta parts per million) with the use of previously published methods. 15 Mierobiologic evaluations. Fecal samples were obtained from each of the patients at the time of the breath H2 studies. Samples were examined for recognized bacterial pathogens by means of standard microbiologic cultivation assays. Stools were examined for parasitic agents by formol concentration and polyvinyl alcohol-stained smears. The latter were prepared in three ways: trichrome stain, au-
The Journal of Pediatrics March 1991
ramine O, and the modified Kinyoun acid fast stain. The slides were examined by ultraviolet or light microscopy as appropriate for the stain.16 Samples were also examined for the presence of group A rotaviruses, group B rotaviruses, and enteric adenoviruses by means of enzyme immunoassays with previously published methods. 17 The fecal samples were also examined for the presence of cytotoxins and cultivatable viruses by inoculation in Vero cells, with the use of standard methods. Intestinal biopsies. Intestinal biopsy samples were obtained from four of the children. Patients were selected for endoscopy and biopsy on the basis of clinical indications as determined by their physicians. The patients were sedated with meperidine and diazepam or underwent general anesthesia. Upper gastrointestinal tract endoscopy was performed with a model G I F - X P fiber-optic endoscope (Olympus Corp. of America, New Hyde Park, N.Y.) Duodenal fluid was obtained by passing polyethylene tubing through the biopsy channel of the endoscope. Endoscopic biopsy specimens were obtained from the distal portion of the duodenum, duodenal bulb, gastric antrum, and esophagus in two patients; the duodenum in a third; and the esophagus in a fourth. Fluid obtained during the endoscopies was transported immediately to the microbiology laboratory and cultured under aerobic and anaerobic conditions. All biopsy specimens were fixed in Hollande solution and processed into 5 #m thick hematoxylin-eosin-stained sections. The samples were also processed with periodic acidSchuff/alcian blue, Giemsa, modified acid-fast, and BrownBrenn stains to detect fungal, bacterial, and protozoal organisms. Hollande-fixed sections were also examined immunohistochemically for the presence of HIV p24 antigen by a method similar to that developed by Ullrich et al. 7 This method consists of the reaction of tissue sections with murine monoclonal anti-p24 antibody (IgG1 subclass, Dupont Laboratories, Wilmington, Del.) and peroxidaselabeled goat antimouse IgG (K & P Laboratories, Gaithersburg, Md.). The reactivity of the antibodies was demonstrated by the staining of HIV antigen in the HIV-infected, 8E5 cell linO 8 with techniques identical to those used for processing the intestinal tissues. Statistical analyses. Quantitative data were analyzed by means of nonparametric methods including the MannWhitney I.I test (two-way). In the case of patients who had more than one breath H2 test for lactose, the first determination was used for the purpose of statistical analysis. Calculations were performed with the aid of the CSS statistical package (Statsoft Inc.; Tulsa, Okla.). RESULTS We studied 17 children with evidence of HIV infection. Nine of these children had clinical manifestations of gastrointestinal dysfunction at the time of initial study as
Volume 118 Number 3
Gastrointestinal dysfunction in H I V infection
361
800
|
100"
t Jo EN Z
W
1 O:
8
II,
0 0 O0O
J
QI Disease
NO GI Disease
Fig. 1. Breath H2 excretion after lactose ingestion. Children in study population were fed lactose after overnight fast, and peak level of breath H2 above baseline level (~ ppm) was measured by gas chromatography as described in text. Clear and dark circles represent peak concentrations measured for HIV-infected children with and without persistent diarrhea, respectively, with definitions given in text. GL Gastrointestinal.
evidenced by persistent diarrhea ranging in duration from 1 to 24 months. The HIV-infected children with and without persistent diarrhea did not differ in terms of age, race, gender, length of time since diagnosis, or duration of administration of 3 '-azido-2',3 '-dideoxythymidine. Breath H2 analyses were performed on all children in the study, after the ingestion of lactose. As depicted in Fig. 1, we found a strong association between persistent diarrheal disease and lactose malabsorption. The geometric mean peak elevation of breath H2 was 49.2 6 ppm (95% confidence limits, 25.1 to 96.5 6 ppm), compared with 4.8 6 ppm (95% confidence limits, 2.3 to 10.3 ~ ppm) for the HIV-infected children without evidence of persistent diarrhea (p <0.007, Mann-Whitney U test). Three patients with gastrointestinal disease were also studied for the production of hydrogen after sucrose administration. 13 All three had elevated breath H2 concentrations after sucrose ingestion (59, 29, and 18 6 ppm, respectively; normal concentration, <10 6 ppm). A total of 9 of the 17 children studied weighed at or below the 5th percentile for age. Six of these children had persistent diarrheal disease; three did not have a history of diarrhea. Upper gastrointestinal endoscopy was performed on four HIV-infected children with gastrointestinal dysfunction. No macroscopic abnormalities were visualized in any of the
children. One patient had histologic evidence of chronic peptic duodenitis with moderate blunting of villi, gastric mucin cell metaplasia, and intramucosal Brunner glands. This same patient had active chronic inflammation at the gastroesophageal junction, which also appeared peptic in origin. A second patient had chronically inflamed duodenal mucosa (Fig. 2) and an acute and chronic inflammation of the distal portion of the esophagus. A third patient had two different biopsies of the duodenum, but neither specimen displayed specific abnormalities. In the fourth patient an esophageal biopsy specimen displayed only minimal reactive changes. None of the samples revealed evidence of bacterial, fungal, or protozoal infection by means of modified acid-fast, periodic acid-Schiff, Giemsa, or Brown-Brenn staining. None of the samples displayed intranuclear inclusions characteristic of infection with cytomegalovirus. The biopsy samples were also examined for the presence of HIV antigen by means of immunoperoxidase staining with monoclonal anti-HIV antibodies; HIV-infected cells could not be detected in any of the biopsy samples. Microscopic analysis of fecal or intestinal fluid samples failed to reveal any infections caused by Cryptosporidium, G. lamblia, or other parasites in any of the study patients. Cultivation analyses revealed no recognized bacterial enteropathogens, cytotoxins, or pathogenic viruses, and immunoassays did not detect any infections caused by rotavi-
362
Yolken et al.
The Journal of Pediatrics March 1991
DISCUSSION
Fig. 2. Intestinal biopsyspecimenfrom 15-month-oldinfant with HIV infection and persistent diarrheal disease. A, Low-powerview of chronically inflamed duodenal mucosa with distorted and blunted villi. (Hematoxylin-eosin stain; original magnification, • B, High-power view of distorted villus showing surface epithelium with increased intraepithelial lymphoeytes.(Hematoxylin-eosin stain; original magnification, •
ruses, adenoviruses, or related agents. One patient had evidence of infection with Mavium-intracellulare as indicated by the persistent isolation of this organism from the blood. One HIV-infected infant was followed for an extended period. When first tested at 12 months of age, he had persistent diarrhea and failure to thrive, and there was evidence of marked lactose malabsorption (peak breath Hz, 137 6 ppm). He was treated as an inpatient with nasogastric feedings with a non-lactose-containing,soy-based formula and had improvement in his symptoms and weight gain. Two months after his initial breath Hz evaluation, he was found to absorb lactose normally (breath Hz, 4 6 ppm). However, after he left the hospital environment, his symptoms returned and at 22 months of age he again had evidence of severe lactose malabsorption (breath Hz, 264 b ppm).
The results of our studies provide documentation that persistent diarrhea and disaccharide malabsorption are common manifestations of HIV infection in children living in the United States. Our results are similar to those of Miller et al., 19 who also found a high incidence of lactose intolerance in infants and young children infected with HIV. In contrast to adults with HIV infection and gastrointestinal dysfunction,57 the children in our study did not have a high rate of infection with recognized opportunistic enteric pathogens, probably because they were not exposed to environmental conditions or to sexual practices that favor the transmission of such pathogens. We also could not document a role for intestinal HIV replication in the pathogenesis of gastrointestinal disease in our patients, as has been described for some HIV-infected adults. 7 However, the size of the biopsy specimens was small, and it is possible that low levels of intestinal HIV infection were not detected by the methods used. Our microbiologic and histologic findings support the hypothesis that HIV disease can be associated with gastrointestinal dysfunction in the absence of direct evidence of enteric infection with recognizable enteropathogenic agents. 7 However, the possible occurrence of opportunistic infections with microsporidia, fastidious viruses, enteropathogenic strains of Escherichia coli, or other pathogens not detectable by these methods cannot be excluded. Additional studies should be directed toward elucidating the role of such agents, as well as diet, food allergies, and other factors, in the pathogenesis of gastrointestinal disease in HIVinfected children. The mechanisms involved in the development of disaccharide malabsorption in the HIV-infected children are not known. It is possible that the disaccharide malabsorption was caused by mucosal injury. Intestinal villous blunting was documented in two patients, both of whom had lactose malabsorption. Moreover, three patients had malabsorption of both lactose and sucrose, suggesting nonspecific effects on the intestinal brush border rather than an isolated enzyme deficiency. Recovery of lactose absorption was noted transiently in one patient studied serially at a time when the child's nutritional status had improved. These initial studies did not include D-xylose absorption assays, which are useful in the detection of mucosal injury,z~ Our findings indicate that, regardless of cause, carbohydrate malabsorption is an important component of HIV infection in children living in the United States. Although we used lactose as the principal substrate for the analyses, our data indicate that children with HIV infection can also have malabsorption of sucrose. Additional studies of the absorption of sucrose, glucose, and other sugars should be per-
Volume 118 Number 3
formed to establish a better definition of the nature of the absorptive defect in children with H I V infection. The malabsorption of disaccharides and other essential nutrients, in addition to leading to growth retardation and developmental delay, might result in immunologic abnormalities that would exacerbate the T cell defects inherent in H I V infection, z Gastrointestinal dysfunction might also adversely affect the ability of HIV-infected children to absorb zidovudine and other orally administered drugs. 21 Although our study was prospective in nature, we were not able to identify and evaluate every HIV-infected child in our population. It is possible that the children whom we studied were not representative of all children with H I V infection but, rather, represented children with more symptomatic disease and thus who were more likely to seek medical attention and to comply with the study regimens. The actual incidence of gastrointestinal dysfunction and disaccharide intolerance in children with H I V infection should thus be the subject of additional studies. Although the cause of gastrointestinal dysfunction in children with H I V infection is multifactorial, it is possible that some of the symptoms are related to carbohydrate malabsorption and are thus amenable to dietary management. In addition to the loss of calories in the form of dietary sugars that are not absorbed, the osmotic diarrheal state further affects nutritional status by leading to frequent restrictions of dietary intake, z2 It is therefore important to identify specific dietary causes of diarrhea and to modify dietary intake accordingly. The alleviation of gastrointestinal symptoms and resulting nutritional deficiencies might reduce the morbidity and ultimately the mortality rate associated with H I V infection in children. REFERENCES
1. Bowen DL, Lane HC, Fauci AS. Immunopathogenesis of the acquired immunodeficiency syndrome. Ann Intern Med 1985; 103:704-9. 2. Falloon J, Eddy J, Wiener L, Pizzo PA. Human immunodeficiency virus infection in children. J PEDIATR 1989;114:1-30. 3. Dworkin B, Wormser GP, Rosenthal WS, et al. Gastrointestinal manifestations of the acquired immunodeficiency syndrome: a review of 22 cases. Am J Gastroenterol 1985;80: 774-8. 4. Kotler DP, Gaetz HP, Lange M, Klein EB, Holt PR. Enteropathy associated with the acquired immunodeficiency syndrome. Ann Intern Med 1984;101:42l-8. 5. Gillin JS, Shike M, Alcock N, et al. Malabsorption and mucosal abnormalities of the small intestine in the acquired immunodeficiency syndrome. Ann Intern Med 1985; 102:619-22.
Gastrointestinal dysfunction in H I V infection
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6. Rene E, Marche C, Regnier B, et al. Intestinal infections in patients with acquired immunodeficiency syndrome: a prospective study in 132 patients. Digest Dis Sci 1989;34: 773-80. 7. Ullrich R, Zeitz M, Heise W, L'age M, Hoffken G, Riecken EO. Small intestinal structure and function in patients infected with human immunodeficiency virus (HIV): evidence for HIV-induced enteropathy. Ann Intern Med 1989;11I: 15-21. 8. Chandra RK. Nutrition, immunity, and infection: present knowledge and future directions. Lancet 1983;1:688-91. 9. Lesbordes JL, Chassignol S, Ray E, et al. Malnutrition and HIV infection in children in the Central Africa Republic. Lancet 1989;2:337-8. 10. McLoughlin LC, Nord KS, Joshi VV, Oleske JM, Connor EM. Severe gastrointestinal involvement in children with the acquired immunodeflciency syndrome. J Pediatr Gastroenterol Nutr 1987;6:517-24. 11. Schuerman L, Seynhaeve V, Bachschmidt I, et al. Severe malnutrition and pediatric AIDS: a diagnostic problem in rural Africa. AIDS 1988;2:232-3. 12. Classification system for human immunodeficiencyvirus (HIV) infection in children under 13 years of age. MMWR 1987; 36:225-36. 13. Perman JA, Barr RG, Watkins JB. Sucrose malabsorption in children: non-invasive diagnosis by interval breath hydrogen determination. J PEDIATR 1978;93:17-23. 14. Perrnan JA, Modler S, Engel RR, Heldt G. Effects of ventilation on breath hydrogen (H2) measurements. J Lab Clin Med 1985;105:426-39. I5. Barr RG, Watkins JB, Perman JA. Mucosal function and breath hydrogen excretion: comparative studies in the clinical evaluation of children with non-specific abdominal complaints. Pediatrics 1981;68:526-33. 16. Ungar BLP, Fayer R. Cryptosporidiurn spp. and cryptosporidiosis. Microbiol Rev 1986;50:458-83. 17. Yolken RH, Viscidi R, Leister F, Harris C, Wee S-B. Enzyme immunoassay for the detection of rotavirus antigen and antibody. In: Rose NR, Friedman H, Fahey JL, eds. Manual of clinical laboratory immunology. Washington, D.C.: American Society for Microbiology, 1986:573-81. 18. Folks TM, Powell D, Lightfoote M, et al. Biological and biochemical characterization of a cloned LeU-3 cell surviving infection with the acquired immunodeficiency syndrome retrovirus. J Exp Med 1988;164:280-90. 19. Miller TL, Martin SR, Cooper ER, Mclntosh K, Winter HS. Gastrointestinal inflammation and carbohydrate intolerance in HIV-infected children [Abstract 650]. Presented at the Annual Meeting of the Society for Pediatric Research, 1990. 20. Christie DL. Use of the one-hour blood xylose test as an indication of small bowel mucosal disease. J PEDIATR 1978;92: 725-8. 21. Pizzo PA. Emerging concepts in the treatment of HIV infection in children. JAMA 1989;262:1989-92. 22. Lloyd-Still JD. Chronic diarrhea of childhood and the misuse of elimination diets. J PEDIATR 1979;95:10-3.
Because gastrointestinal dysfunction is a major problem in children with human immunodeficiency virus (HIV) infection, we utilized breath hydrogen measurements to determine the relationship between disaccharide malabsorption and gastrointestinal dysfunction in HIV-infected children. We found a strong association between lactose intolerance and persistent diarrheal disease in this population (p <0.007, Mann-Whitney U test). We also found evidence of sucrose malabsorption and persistent diarrheal disease in three of the children. Extensive microbiologic evaluations failed to reveal an etiologic agent related to the occurrence of gastrointestinal symptoms. Our findings indicate that disaccharide intolerance is a common occurrence in HIV-infected children with persistent diarrheal disease. Careful attention to dietary intake may be required to ameliorate clinical symptoms and to maintain a d e q u a t e nutrition. (J PEDIATR 1991;118:359-63)
Human immunodeficiency virus replicates in the lymphocytes and macrophages of infected persons; many of the clinical manifestations of HIV infection are due to immune cell dysfunction and subsequent immunodeficiency. 1 However, it is being increasingly recognized that HIV infection results in a multisystem disorder and that dysfunction of other organ systems can contribute to the disease state. 2 One such organ system is the gastrointestinal tract. Acute and chronic diarrhea, weight loss, abdominal pain, esophagitis, and other gastrointestinal symptoms have been noted in a substantial portion of adults with HIV infection. 35 In adults these symptoms are often, but not invariably,
Supported by grant R01-DK-40540 from the National Institutes of Health. Submitted for publication Aug. 24, 1990; accepted Oct. 1, 1990. Reprint requests: Robert H. Y01ken, MD, Pediatric Infectious Diseases, CMSC 1109, Johns Hopkins Hospital, Baltimore, MD 21205. 9/20/25781
associated with infections with gastrointestinal pathogens such as Cryptosporidium, Giardia lamblia, Mycobacteriurn avium-intracellulare, cytomegalovirus, and other agents.6, 7 Gastrointestinal dysfunction can predispose infants and young children to electrolyte imbalance and malnutrition, which may further exacerbate the T cell defects associated with HIV replication.8, 9 Furthermore, the reliHIV
Human immunodeficiency virus
[
ance of infants on milk products as the principal source of nutrition might make infants particularly susceptible to the effects of disaccharidase deficiency and other absorptive abnormalities related to epithelial cell damage. Severe gastrointestinal dysfunction and malnutrition have been reported in children with HIV infection, 1~ 11 but the mechanisms involved have not been elucidated. We utilized breath hydrogen measurements to evaluate carbohydrate absorption prospectively in HIV-infected infants and children.
359
360
Yolken et al.
METHODS Patient population. The study population consisted of 17 infants with documented HIV infection who were followed at Johns Hopkins Hospital and Mount Washington Pediatric Hospital, both located in Baltimore, Md. All children met the criteria for HIV infection in children as defined by persistently positive antibody titers for HIV for longer than 15 months (as documented by enzyme-linked immunosorbent assay and Western blot analyses), or, in the cas~of younger children, by an acquired immunodeficieney syndrome-defining illness or by the identification of HIV in peripheral blood lymphocytes. 12 Patients ranged in age from 4 to 80 months. Seven of the children were receiving 3'-azido-2',3'-dideoxythymidine at the time of analysis. Clinical data were collected from the patients by standard methods. Patients were judged to have persistent diarrheal disease if they had more than three watery bowel movements per day for a minimum of 2 weeks. Informed consent was obtained from the parents or legal guardians of all study subjects. The study protocols were approved by the Johns Hopkins School of Medicine Joint Committee on Human Experimentation. Breath hydrogen analyses. Breath H2 was measured after oral lactose administration with the use of previously described methods. 13 Patients fasted for either 6 hours, in the case of infants less than 1 year of age, or 8 hours in the case of older children. The children were fed lactose at a dosage of 2 g m / k g (maximum50 gm) in the form of a 20% aqueous lactose solution. Three of the patients were also examined for breath H2 generated after the oral administration of 20% aqueous solutions of sucrose at a dosage of 2 gm/kg. Breath sample s were collected by the nasal prong technique at 0, 30, 60, 90, 120, 150, and 180 minutes after disaccharide ingestion. 13 Breath samples were stored in 30 ml syringes for analysis on the day of collection, or in specialized 20 ml evacuated rubber-stoppered glass tubes if analysis was deferred to a subsequent day. Hydrogen concentration was determined in parts per million by gas chromatography at 25 ~ C with a Quintron detector (Quintron Instruments, Milwaukee, Wis.). Each result was corrected to an alveolar H2 concentration with carbon dioxide as an internal standard. 14 The results of each assay were expressed in terms of the peak corrected H2 concentration above the baseline value (delta parts per million) with the use of previously published methods. 15 Mierobiologic evaluations. Fecal samples were obtained from each of the patients at the time of the breath H2 studies. Samples were examined for recognized bacterial pathogens by means of standard microbiologic cultivation assays. Stools were examined for parasitic agents by formol concentration and polyvinyl alcohol-stained smears. The latter were prepared in three ways: trichrome stain, au-
The Journal of Pediatrics March 1991
ramine O, and the modified Kinyoun acid fast stain. The slides were examined by ultraviolet or light microscopy as appropriate for the stain.16 Samples were also examined for the presence of group A rotaviruses, group B rotaviruses, and enteric adenoviruses by means of enzyme immunoassays with previously published methods. 17 The fecal samples were also examined for the presence of cytotoxins and cultivatable viruses by inoculation in Vero cells, with the use of standard methods. Intestinal biopsies. Intestinal biopsy samples were obtained from four of the children. Patients were selected for endoscopy and biopsy on the basis of clinical indications as determined by their physicians. The patients were sedated with meperidine and diazepam or underwent general anesthesia. Upper gastrointestinal tract endoscopy was performed with a model G I F - X P fiber-optic endoscope (Olympus Corp. of America, New Hyde Park, N.Y.) Duodenal fluid was obtained by passing polyethylene tubing through the biopsy channel of the endoscope. Endoscopic biopsy specimens were obtained from the distal portion of the duodenum, duodenal bulb, gastric antrum, and esophagus in two patients; the duodenum in a third; and the esophagus in a fourth. Fluid obtained during the endoscopies was transported immediately to the microbiology laboratory and cultured under aerobic and anaerobic conditions. All biopsy specimens were fixed in Hollande solution and processed into 5 #m thick hematoxylin-eosin-stained sections. The samples were also processed with periodic acidSchuff/alcian blue, Giemsa, modified acid-fast, and BrownBrenn stains to detect fungal, bacterial, and protozoal organisms. Hollande-fixed sections were also examined immunohistochemically for the presence of HIV p24 antigen by a method similar to that developed by Ullrich et al. 7 This method consists of the reaction of tissue sections with murine monoclonal anti-p24 antibody (IgG1 subclass, Dupont Laboratories, Wilmington, Del.) and peroxidaselabeled goat antimouse IgG (K & P Laboratories, Gaithersburg, Md.). The reactivity of the antibodies was demonstrated by the staining of HIV antigen in the HIV-infected, 8E5 cell linO 8 with techniques identical to those used for processing the intestinal tissues. Statistical analyses. Quantitative data were analyzed by means of nonparametric methods including the MannWhitney I.I test (two-way). In the case of patients who had more than one breath H2 test for lactose, the first determination was used for the purpose of statistical analysis. Calculations were performed with the aid of the CSS statistical package (Statsoft Inc.; Tulsa, Okla.). RESULTS We studied 17 children with evidence of HIV infection. Nine of these children had clinical manifestations of gastrointestinal dysfunction at the time of initial study as
Volume 118 Number 3
Gastrointestinal dysfunction in H I V infection
361
800
|
100"
t Jo EN Z
W
1 O:
8
II,
0 0 O0O
J
QI Disease
NO GI Disease
Fig. 1. Breath H2 excretion after lactose ingestion. Children in study population were fed lactose after overnight fast, and peak level of breath H2 above baseline level (~ ppm) was measured by gas chromatography as described in text. Clear and dark circles represent peak concentrations measured for HIV-infected children with and without persistent diarrhea, respectively, with definitions given in text. GL Gastrointestinal.
evidenced by persistent diarrhea ranging in duration from 1 to 24 months. The HIV-infected children with and without persistent diarrhea did not differ in terms of age, race, gender, length of time since diagnosis, or duration of administration of 3 '-azido-2',3 '-dideoxythymidine. Breath H2 analyses were performed on all children in the study, after the ingestion of lactose. As depicted in Fig. 1, we found a strong association between persistent diarrheal disease and lactose malabsorption. The geometric mean peak elevation of breath H2 was 49.2 6 ppm (95% confidence limits, 25.1 to 96.5 6 ppm), compared with 4.8 6 ppm (95% confidence limits, 2.3 to 10.3 ~ ppm) for the HIV-infected children without evidence of persistent diarrhea (p <0.007, Mann-Whitney U test). Three patients with gastrointestinal disease were also studied for the production of hydrogen after sucrose administration. 13 All three had elevated breath H2 concentrations after sucrose ingestion (59, 29, and 18 6 ppm, respectively; normal concentration, <10 6 ppm). A total of 9 of the 17 children studied weighed at or below the 5th percentile for age. Six of these children had persistent diarrheal disease; three did not have a history of diarrhea. Upper gastrointestinal endoscopy was performed on four HIV-infected children with gastrointestinal dysfunction. No macroscopic abnormalities were visualized in any of the
children. One patient had histologic evidence of chronic peptic duodenitis with moderate blunting of villi, gastric mucin cell metaplasia, and intramucosal Brunner glands. This same patient had active chronic inflammation at the gastroesophageal junction, which also appeared peptic in origin. A second patient had chronically inflamed duodenal mucosa (Fig. 2) and an acute and chronic inflammation of the distal portion of the esophagus. A third patient had two different biopsies of the duodenum, but neither specimen displayed specific abnormalities. In the fourth patient an esophageal biopsy specimen displayed only minimal reactive changes. None of the samples revealed evidence of bacterial, fungal, or protozoal infection by means of modified acid-fast, periodic acid-Schiff, Giemsa, or Brown-Brenn staining. None of the samples displayed intranuclear inclusions characteristic of infection with cytomegalovirus. The biopsy samples were also examined for the presence of HIV antigen by means of immunoperoxidase staining with monoclonal anti-HIV antibodies; HIV-infected cells could not be detected in any of the biopsy samples. Microscopic analysis of fecal or intestinal fluid samples failed to reveal any infections caused by Cryptosporidium, G. lamblia, or other parasites in any of the study patients. Cultivation analyses revealed no recognized bacterial enteropathogens, cytotoxins, or pathogenic viruses, and immunoassays did not detect any infections caused by rotavi-
362
Yolken et al.
The Journal of Pediatrics March 1991
DISCUSSION
Fig. 2. Intestinal biopsyspecimenfrom 15-month-oldinfant with HIV infection and persistent diarrheal disease. A, Low-powerview of chronically inflamed duodenal mucosa with distorted and blunted villi. (Hematoxylin-eosin stain; original magnification, • B, High-power view of distorted villus showing surface epithelium with increased intraepithelial lymphoeytes.(Hematoxylin-eosin stain; original magnification, •
ruses, adenoviruses, or related agents. One patient had evidence of infection with Mavium-intracellulare as indicated by the persistent isolation of this organism from the blood. One HIV-infected infant was followed for an extended period. When first tested at 12 months of age, he had persistent diarrhea and failure to thrive, and there was evidence of marked lactose malabsorption (peak breath Hz, 137 6 ppm). He was treated as an inpatient with nasogastric feedings with a non-lactose-containing,soy-based formula and had improvement in his symptoms and weight gain. Two months after his initial breath Hz evaluation, he was found to absorb lactose normally (breath Hz, 4 6 ppm). However, after he left the hospital environment, his symptoms returned and at 22 months of age he again had evidence of severe lactose malabsorption (breath Hz, 264 b ppm).
The results of our studies provide documentation that persistent diarrhea and disaccharide malabsorption are common manifestations of HIV infection in children living in the United States. Our results are similar to those of Miller et al., 19 who also found a high incidence of lactose intolerance in infants and young children infected with HIV. In contrast to adults with HIV infection and gastrointestinal dysfunction,57 the children in our study did not have a high rate of infection with recognized opportunistic enteric pathogens, probably because they were not exposed to environmental conditions or to sexual practices that favor the transmission of such pathogens. We also could not document a role for intestinal HIV replication in the pathogenesis of gastrointestinal disease in our patients, as has been described for some HIV-infected adults. 7 However, the size of the biopsy specimens was small, and it is possible that low levels of intestinal HIV infection were not detected by the methods used. Our microbiologic and histologic findings support the hypothesis that HIV disease can be associated with gastrointestinal dysfunction in the absence of direct evidence of enteric infection with recognizable enteropathogenic agents. 7 However, the possible occurrence of opportunistic infections with microsporidia, fastidious viruses, enteropathogenic strains of Escherichia coli, or other pathogens not detectable by these methods cannot be excluded. Additional studies should be directed toward elucidating the role of such agents, as well as diet, food allergies, and other factors, in the pathogenesis of gastrointestinal disease in HIVinfected children. The mechanisms involved in the development of disaccharide malabsorption in the HIV-infected children are not known. It is possible that the disaccharide malabsorption was caused by mucosal injury. Intestinal villous blunting was documented in two patients, both of whom had lactose malabsorption. Moreover, three patients had malabsorption of both lactose and sucrose, suggesting nonspecific effects on the intestinal brush border rather than an isolated enzyme deficiency. Recovery of lactose absorption was noted transiently in one patient studied serially at a time when the child's nutritional status had improved. These initial studies did not include D-xylose absorption assays, which are useful in the detection of mucosal injury,z~ Our findings indicate that, regardless of cause, carbohydrate malabsorption is an important component of HIV infection in children living in the United States. Although we used lactose as the principal substrate for the analyses, our data indicate that children with HIV infection can also have malabsorption of sucrose. Additional studies of the absorption of sucrose, glucose, and other sugars should be per-
Volume 118 Number 3
formed to establish a better definition of the nature of the absorptive defect in children with H I V infection. The malabsorption of disaccharides and other essential nutrients, in addition to leading to growth retardation and developmental delay, might result in immunologic abnormalities that would exacerbate the T cell defects inherent in H I V infection, z Gastrointestinal dysfunction might also adversely affect the ability of HIV-infected children to absorb zidovudine and other orally administered drugs. 21 Although our study was prospective in nature, we were not able to identify and evaluate every HIV-infected child in our population. It is possible that the children whom we studied were not representative of all children with H I V infection but, rather, represented children with more symptomatic disease and thus who were more likely to seek medical attention and to comply with the study regimens. The actual incidence of gastrointestinal dysfunction and disaccharide intolerance in children with H I V infection should thus be the subject of additional studies. Although the cause of gastrointestinal dysfunction in children with H I V infection is multifactorial, it is possible that some of the symptoms are related to carbohydrate malabsorption and are thus amenable to dietary management. In addition to the loss of calories in the form of dietary sugars that are not absorbed, the osmotic diarrheal state further affects nutritional status by leading to frequent restrictions of dietary intake, z2 It is therefore important to identify specific dietary causes of diarrhea and to modify dietary intake accordingly. The alleviation of gastrointestinal symptoms and resulting nutritional deficiencies might reduce the morbidity and ultimately the mortality rate associated with H I V infection in children. REFERENCES
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