RESUMO
OBJECTIVE: Epifluorescence microscopy, a methodology for the screening of bodily fluids and tissue specimens for microsporidia species, was directed to evaluate the retention of epifluorescence of fixed and stained specimens over time. METHODS: Thirty samples of stool, bodily fluids, duodenal touch preparations, and biopsies, were tested for the retention of their epifluoresence using the Fungi-Fluor procedure. Specimens were examined under a 330- to 380-nm UV filter at the time of preparation, 3 wk later, and then at monthly intervals for 18 months. All specimens were reevaluated for the presence or absence of fluorescence and any decrement of fluorescence over time. No special preservation techniques were used on any of the slides. RESULTS: All 30 specimens maintained their epifluorescence from the time of slide preparation to 18 month later. No decrement in fluorescence was noted in any sample examined. Accuracy and ease of spore identification was maintained. CONCLUSIONS: Epifluorescence microscopy demonstrates the utility of this technique for archival study of microsporidia-containing specimens over prolonged periods of time.
Assuntos
Corantes Fluorescentes , Microsporídios/isolamento & purificação , Animais , Corantes , Fluorescência , Humanos , Microscopia de Fluorescência , Fatores de TempoRESUMO
Gastrointestinal microsporidiosis is a major cause of diarrhea and wasting in persons with acquired immune deficiency syndrome (AIDS). Microsporidia demonstrate properties of both true eukaryotes and prokaryotes. The biology of microsporidia makes its elimination from the gastrointestinal tract therapeutically challenging. This organism depends greatly on the host for its energy needs and reproduction; microsporidial spores are impervious to the elements. Microsporidial infection of the gastrointestinal tract, principally with Enterocytozoon bieneusi and Encephalitozoon intestinalis in patients with AIDS has been treated with different medical regimens with variable success. The less common pathogen, E. intestinalis, responds well to albendazole, making it excellent first-line therapy, but such is not the case for E. bieneusi. None of the benzimidazoles has been demonstrated to be efficacious for E. bieneusi. On the other hand, E. bieneusi has shown excellent clinical therapeutic response to either direct action with fumagillin or its analogue, TNP-470, or indirectly by immune enhancement by suppression of the HIV virus with more aggressive, highly effective antiretroviral therapy. Further work is necessary to fully establish proper therapeutic protocols and manage side effects of the treatments. Other promising forms of therapy such as polyamine inhibitors and thalidomide demonstrate certain effectiveness in treatment of microsporidian in vitro (polyamine inhibitors) and in selected cases in vivo (thalidomide). Lack of either sufficiently suggestive or definitive human studies prevents the endorsement of these modes of therapy for treatment of gastrointestinal microsporidiosis at this time.
Assuntos
Infecções Oportunistas Relacionadas com a AIDS/tratamento farmacológico , Antiprotozoários/uso terapêutico , Enteropatias Parasitárias/tratamento farmacológico , Microsporidiose/tratamento farmacológico , HumanosRESUMO
The clinical course of 37 Enterocytozoon bieneusi-infected acquired immunodeficiency syndrome patients with diarrhea was studied. Parasite clearance was seen in 15 patients (40.5%). Clearance of E. bieneusi resulted in a 25-100% reduction in episodes of diarrhea, suggesting that microsporidia are true pathogens. Univariate and multivariate proportional hazards analyses revealed that peripheral blood CD4 cell counts > or = 100/mm3, the use of two or more antiretroviral medications, and use of a protease inhibitor were statistically associated with decreased time to clearance of E. bieneusi. Specific anti-microsporidial therapy (albendazole) was not associated with parasite eradication. Factors related to immunocompetence and human immunodeficiency virus suppression appeared to be important in the clearance of E. bieneusi.
Assuntos
Infecções Oportunistas Relacionadas com a AIDS/parasitologia , Enteropatias Parasitárias/parasitologia , Microsporidiose/parasitologia , RNA Viral/sangue , Infecções Oportunistas Relacionadas com a AIDS/sangue , Síndrome da Imunodeficiência Adquirida/tratamento farmacológico , Adulto , Análise de Variância , Animais , Fármacos Anti-HIV/uso terapêutico , Humanos , Hospedeiro Imunocomprometido , Enteropatias Parasitárias/etiologia , Masculino , Microsporida/isolamento & purificação , Microsporidiose/etiologia , Pessoa de Meia-Idade , Inibidores de Proteases/uso terapêuticoRESUMO
The epidemiology of human microsporidiosis is poorly understood and environmental factors affecting transmission of the organism have not been fully elucidated. Temporal variation in the prevalence of microsporidia in the stool of patients with human immunodeficiency virus (HIV) infection and diarrhea was studied to evaluate the role of water-borne transmission. From January 1993 to December 1996, 8,439 stools from HIV-infected individuals were examined for microsporidia spores in southern California. Yearly positivity rates were 8.8% in 1993, 9.7% in 1994, 6.6% in 1995, and 2.9% in 1996. An analysis for linear trend showed a statistically significant decrease in stool positivity rates over time (chi2 = 81.9, P = 0.001). No significant seasonal variation in the prevalence of microsporidiosis was seen over that time period. These results suggest the constant presence of microsporidia in the environment, rather than a seasonal association with recreational water use or seasonal contamination of the water supply, and a real decrease in yearly prevalence of microsporidia related diarrhea. Factors related to a progressive decrease in prevalence are subjects of future investigation.
Assuntos
Diarreia/parasitologia , Infecções por HIV/complicações , Enteropatias Parasitárias/epidemiologia , Microsporidiose/epidemiologia , Animais , Doença Crônica , Fezes/parasitologia , Infecções por HIV/epidemiologia , Humanos , Umidade , Enteropatias Parasitárias/parasitologia , Microsporida/isolamento & purificação , Microsporidiose/parasitologia , Prevalência , Estações do AnoRESUMO
Gastrointestinal ulcerations in persons infected with HIV have many causes, the most common being opportunistic infections and neoplasms. Recently, idiopathic ulcerative lesions of the colon and rectum have been described. Two cases are reported of idiopathic colonic and anorectal inflammation and ulceration which failed traditional therapies but responded to thalidomide with complete clinical and histologic resolution.
Assuntos
Colite Ulcerativa/tratamento farmacológico , Infecções por HIV/complicações , Imunossupressores/uso terapêutico , Proctite/tratamento farmacológico , Talidomida/uso terapêutico , Adulto , Colite Ulcerativa/complicações , Humanos , Masculino , Pessoa de Meia-Idade , Proctite/complicaçõesRESUMO
Microsporidia, which are members of the phylum Microspora, are increasingly recognized as causing opportunistic infections in persons with immunodeficiency (e.g., AIDS). Diarrhea is the predominant clinical sign associated with infections by two Microsporidia, namely Enterocytozoon bieneusi and Encephalitozoon intestinalis (which was formerly named Septata intestinalis). Prevalence rates of microsporidiosis in persons with AIDS and chronic diarrhea range fron 7 to 50%. Transmission electron microscopy has been the gold standard by which to diagnose microsporidiosis and requires observing a polar filament which is the structure distinguishing Microsporidia from other organisms. Transmission electron microscopy is difficult, time-consuming, costly, relatively insensitive, and requires a great deal of expertise. As such, histochemical methods have been developed and improved for detecting Microsporidia. Diagnoses from stool specimens or enteric fluids can be made using the chitin-staining fluorochromes (e.g., Calcofluor White) and the modified trichrome stain which are highly sensitive, particularly when both are used. Immunofluorescent antibody staining methods are being developed to improve specificity, but reagents are not yet commercially available. Microsporidia can be detected most readily in tissue biopsies by Gram stain, Giemsa stain, or immunofluorescent antibody. Polymerase chain reaction methods are in their infancy for application, but should prove to be particularly sensitive and specific in the future.
Assuntos
Gastroenteropatias/diagnóstico , Gastroenteropatias/parasitologia , Microsporidiose , Microsporidiose/diagnóstico , Encefalitozoonose/diagnóstico , Gastroenteropatias/patologia , Humanos , Microsporidiose/patologia , Reação em Cadeia da PolimeraseRESUMO
OBJECTIVE: To evaluate three fluorescent chitin stains for detecting microsporidia spores in specimens from acquired immunodeficiency syndrome (AIDS) patients with chronic diarrhea. METHODS: We compared the Fungifluor, Calcofluor White, and Fungiqual A fluorochrome stains for identifying Enterocytozoon bieneusi and Septata intestinalis spores in stool, intestinal fluid, biopsy imprints, and paraffin biopsy sections. The modified chromotrope trichrome stain was used as the standard light microscopic technique for stool and fluid specimens. Stained and unstained paraffin sections and fluid preparations were also evaluated. Multiple specimens from 50 consecutive symptomatic AIDS patients and archival material from known microsporidia-positive AIDS patients were analyzed. RESULTS: Spores of E bieneusi and S intestinalis fluoresce brightly with all three fluorochrome stains in all of the types of diagnostic specimens. Fluorescing debris and the much larger fungal forms were readily distinguished. Spores were equally well detected in unfixed and formalin-fixed stool specimens, but were not as well detected after sodium acetate-acetic acid, polyvinyl acetate, and ethanol fixation. Bouin's tissue fixative gave a higher background staining than formalin. Spores were readily detected in archival paraffin sections and stool preparations, even when the specimens had been stained previously. Repeat fluorochrome staining was possible. The methods also could detect extraintestinal parasites in paraffin sections. CONCLUSION: The three fluorescent chitin stains are sensitive and rapid methods for detecting microsporidia spores in stool, intestinal fluid, biopsy imprint, and tissue specimens, even from archived material.
Assuntos
Síndrome da Imunodeficiência Adquirida/parasitologia , Diarreia/parasitologia , Corantes Fluorescentes , Microsporida/isolamento & purificação , Microsporidiose/parasitologia , Síndrome da Imunodeficiência Adquirida/complicações , Animais , Benzenossulfonatos , Biópsia , Líquidos Corporais/parasitologia , Quitina/análise , Diarreia/complicações , Fezes/parasitologia , Fixadores , Humanos , Intestinos/parasitologia , Microscopia de Fluorescência , Microsporidiose/complicações , Projetos Piloto , Estilbenos , TriazinasRESUMO
OBJECTIVE: To determine the clinical and endoscopic response of candida esophagitis to antifungal therapy and to compare the two oral antifungal agents, fluconazole and ketoconazole. DESIGN: Multicenter, randomized, double-blind trial. SETTING: Fifteen U.S. centers including university, private practice, and county hospital settings. PATIENTS: A total of 169 patients with the acquired immunodeficiency syndrome (AIDS); odynophagia, dysphagia, or retrosternal pain; white esophageal plaques at endoscopy; and pseudohyphae on esophageal brushings or biopsies. INTERVENTION: Patients were randomly assigned to fluconazole (100 mg/d) or ketoconazole (200 mg/d). Doses were doubled at week 1 or 2 if no symptomatic improvement had occurred during the preceding week. Therapy was continued for 2 weeks after resolution of symptoms or for a maximum of 8 weeks. MEASUREMENTS: Patients were clinically evaluated weekly, and laboratory tests were done every 2 weeks. Endoscopy was repeated within 5 days after the end of therapy. RESULTS: A total of 143 patients were clinically evaluable (assessed within 7 days after therapy), and 129 patients were endoscopically evaluable (endoscopy repeated after therapy). Endoscopic cure occurred in 91% of patients treated with fluconazole and in 52% of those given ketoconazole for a difference of 39% (95% Cl, 24% to 52%; P less than 0.001). Esophageal symptoms resolved in 85% of fluconazole-treated patients and in 65% of ketoconazole-treated patients for a difference of 20% (Cl, 6% to 34%; P = 0.006). Intention-to-treat analyses also yielded statistically significant differences for the comparisons listed above. Side effects were minimal and comparable in the two groups; only one patient in each group had therapy discontinued for adverse effects that were possibly related to the study medications. CONCLUSIONS: Fluconazole is associated with significantly greater rates of endoscopic and clinical cure than ketoconazole in patients with AIDS and candida esophagitis. Both drugs appear to be safe and well tolerated.
Assuntos
Síndrome da Imunodeficiência Adquirida/complicações , Candidíase/tratamento farmacológico , Esofagite/tratamento farmacológico , Fluconazol/uso terapêutico , Cetoconazol/uso terapêutico , Adulto , Candidíase/etiologia , Método Duplo-Cego , Esofagite/microbiologia , Esofagite/patologia , Esofagoscopia , Feminino , Humanos , MasculinoRESUMO
Thymosin alpha 1 (T alpha 1), the N-terminal 28-amino acid fragment of prothymosin alpha (ProT alpha), and ProT alpha, although originally isolated from whole thymus extracts, are also present in nonthymic cells and tissues. We used an ELISA with an antibody raised against T alpha 1 to investigate the relationship between intracellular levels of thymosin immunoreactive peptide(s) (TIP) and cell proliferation in a rat small intestinal IEC-6 cell line. Increasing TIP levels were observed during cell proliferation, which decreased when proliferation was halted by cellular contact inhibition. Serum feeding of cells previously rendered quiescent by serum starvation resulted in a significant increase in TIP within 1 hr. Conversely, serum starvation decreased TIP levels within 1 hr. Peak TIP levels appeared after 3 hr of serum incubation, while maximum [3H]thymidine incorporation was noted after 9 hr, suggesting maximum TIP concentrations in the G1 phase of the proliferative cycle. Immunoelectron microscopy demonstrated an association of TIP with condensed nuclear chromatin. These results support a relation of intracellular TIP levels to IEC-6 cell proliferation and also a nuclear site of action. HPLC analysis of cellular homogenates from proliferating IEC-6 cells revealed a peak of immune reactivity that elutes in the position of T alpha 1.
Assuntos
Divisão Celular , Núcleo Celular/ultraestrutura , Precursores de Proteínas/análise , Timosina/análogos & derivados , Timosina/análise , Animais , Linhagem Celular , Núcleo Celular/fisiologia , Cromatografia Líquida de Alta Pressão , Ensaio de Imunoadsorção Enzimática , Intestino Delgado , Cinética , Microscopia Eletrônica , Radioimunoensaio , Ratos , Ratos Endogâmicos , TimalfasinaRESUMO
The effect of insulin on EGF-induced cell proliferation and on modulation of EGF receptors was examined in IEC-6 cells in culture. Insulin-induced cell proliferation was not seen until concentration of the hormone reached the microgram (10 micrograms/ml) level, producing an increase in EGF receptor binding affinity (Ka 8.2 +/- .05 x 10(9)M-1 to 1.25 +/- .05 10(10) M-1). In the presence of supraphysiologic concentrations of insulin (10 micrograms/ml), EGF-induced cell proliferation could only be detected at a concentration of 50 ng/ml as opposed to a concentration of 2.0 ng/ml in the absence of insulin. This study suggests that insulin stimulates intestinal crypt cell proliferation only in supraphysiologic concentrations in a manner more characteristic of IGF-I than insulin, while producing enhanced binding of EGF to its receptor as well as an elevated threshold to EGF induced cell proliferation.
Assuntos
Divisão Celular/efeitos dos fármacos , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/fisiologia , Insulina/farmacologia , Mucosa Intestinal/citologia , Linhagem Celular , Células Cultivadas , Relação Dose-Resposta a Droga , Técnicas In Vitro , CinéticaRESUMO
The role of hormones and growth factors in the pathogenesis and therapy of colon cancer is biologically intricate and medically important. The effects of the previously described hormones and growth factors on normal and neoplastic colonic growth and development suggest the mechanisms by which hormonal alteration might either enhance or suppress the cancer process. The high degree of association between the specific endocrine-related processes (breast cancer, acromegaly, hyperparathyroidism, gastrin sensitivity of colon cancer, and cancer cell lines) suggests a significant role for hormones in colonic carcinogenesis. The relationship between the specific hormones and cancers is often unclear. This is the result of many factors: the variable presence of specific hormone receptors on the surface of the tumor or cell line; the inconsistent response to exogeneous hormone administration in vivo and in vitro; and the occasional failure of specific hormone-blocking agents to affect cell proliferation. The relationship between growth factors and cancers is also unclear. The following questions must be resolved in order to understand the significance of growth factors and the neoplastic process: (1) Is a growth factor significant in either an autocrine or a paracrine capacity? (2) Are combinations of growth factors rather than individual growth factors more biologically significant? (3) Do structural alterations of the immunologically similar, but functionally different growth factors modify their effect on the neoplastic tissue? The potential for manipulation of hormones and growth factors in the prevention and treatment of colon cancer is evidence to date suggesting that such efforts are indeed justified.
Assuntos
Neoplasias do Colo/fisiopatologia , Substâncias de Crescimento/fisiologia , Hormônios/fisiologia , HumanosRESUMO
Markers that are now in use, including CEA and CA-19-9, are not specific or sensitive enough to detect early colorectal cancer. Newer tumor markers such as polyamines, ornithine decarboxylase, and altered blood group carbohydrate antigens may have a potential as future tumor markers. Additional studies of these markers as well as the development of new biochemical markers are warranted in the future to enhance the sensitivity and specificity of diagnosis of early colorectal cancer and those at risk for developing cancer. Finally, understanding events involved in abnormal cell proliferation (that is, elevated polyamines and ODC in colorectal cancer) may help direct future chemotherapy and possibly chemoprevention in high-risk groups such as adenomatous polyposis coli.
Assuntos
Biomarcadores Tumorais , Neoplasias Colorretais/diagnóstico , Anticorpos Monoclonais , Antígeno Carcinoembrionário , Neoplasias Colorretais/terapia , Humanos , Ornitina DescarboxilaseRESUMO
Exposure of IEC-6 cells for 24 hr to either gastrin (50-500 ng/ml) or EGF (100-500 ng/ml) significantly stimulated (100-165%) the rate of [3H]thymidine incorporation into DNA (referred to as DNA synthesis) when compared with the corresponding basal levels. Somatostatin (10-500 ng/ml) produced no apparent change in DNA synthesis in IEC cells. On the other hand, somatostatin completely inhibited the EGF-induced rise in DNA synthesis. The gastrin-mediated stimulation in DNA synthesis was not affected by somatostatin. The rate of DNA synthesis in IEC cells in the presence of both gastrin and EGF was found to be greater (additive) than that caused by either of the peptides alone. A similar but less dramatic change in the actual number of cells (assessment of cell replication) was observed when the IEC cells were exposed for 24 hr to gastrin, EGF, and somatostatin, either alone or in combination. Whereas gastrin (250 ng/ml) and EGF (250 ng/ml) by themselves increased the number of cells significantly by 29 and 37%, respectively, together they caused a 72% stimulation, when compared with the basal levels. Somatostatin by itself caused no apparent change in IEC cell population, but it significantly inhibited the EGF- but not the gastrin-induced stimulation in IEC cell replication. It is concluded that both gastrin and EGF exert a direct proliferative effect on IEC cells, and the EGF action is regulated by somatostatin.
Assuntos
DNA/biossíntese , Fator de Crescimento Epidérmico/farmacologia , Gastrinas/farmacologia , Intestino Delgado/efeitos dos fármacos , Somatostatina/farmacologia , Animais , Linhagem Celular , Intestino Delgado/citologia , Ratos , Ratos EndogâmicosRESUMO
Circulating concentrations of digestive enzymes, certain lysosomal hydrolases and protease inhibitors were measured in 19 heavy smokers and 13 non-smokers before (basal) and at 15, 30, and 60 minutes after a single intravenous injection of secretin (75 CU). In smokers, basal serum amylase and immunoreactive pancreatic elastase 2 (IRE2) concentrations were about 100% and 25% higher respectively, than in the non-smokers, whereas, no differences were observed in basal immunoreactive cationic trypsinogen (IRCT) concentrations and in acid phosphatase and beta-glucuronidase activities between the two groups. Furthermore, a single injection of secretin to cigarette smokers significantly increased serum amylase, IRCT and IRE2 by 155%, 200%, and 100%, respectively when compared with their corresponding basal levels. No such increment was observed in the non-smokers. In addition, there were no significant differences in serum trypsin or elastase inhibitory capacity or immunoreactive alpha 1-protease inhibitor and alpha 2-macroglobulin levels between smokers and non-smokers. The levels and inhibitory capacity of these protease inhibitors was also not affected by secretin injection. These data suggest that cigarette smoking enhances the responsiveness of the exocrine pancreas to a physiological stimulus such as secretin, with resultant substantial increase in the concentrations of pancreatic hydrolases in blood.