Colitis seudomembranosa como hallazgo posmortemen paciente pediátrico. Reporte de caso / Pseudomembranous colitis as postmortem finding in pediatric patient. Case report

La colitis seudomembranosa es una severa y a veces mortal afección que puede ocurrir tras la administración de antibióticos y supresión de la flora intestinal normal, seguida de colonización por Clostridium difficile; se caracteriza por la inflamación aguda y presencia de seudomembranas necróticas en la mucosa colónica. Se presenta el caso de un paciente varón de nueve años de edad, proveniente de una zona rural de Honduras, con antecedente de fiebre intermitente de cuatro semanas de evolución, escalofríos, mialgias, náuseas e ictericia de una semana de evolución. Atendido previamente en Centro de Atención Primaria, fue tratado con antipiréticos, sin mejoría. Al examen físico el paciente estaba lúcido, se halló hipotensión, taquicardia, y fiebre; dolor abdominal epigastrio y ambos hipocondrios a la palpación superficial y profunda, hepatomegalia, ictericia, petequias. En los exámenes de laboratorio se encontraron pancitopenia severa, falla renal aguda, trastornos hidroelectrolíticos e hipoalbuminemia. Fue ingresado al servicio de urgencias pediátricas. Luego de una mala evolución clínica, falleció diecinueve después del ingreso. La autopsia reveló seudomembranas necróticas colónicas e imagen histológica de tipo volcán compatibles con colitis seudomembranosa.

Pseudomembranous colitis is a severe and often fatal condition that may occur after the administration of some antimicrobial agents. There is suppression of the normal intestinal flora, followed by colonization by Clostridium difficile; and this condition is characterized by acute inflammation and presence of necrotic tissue pseudomembranes in the colon mucosa. We present the case of a nine-year-old boy from a rural area in Honduras, with a history of intermittent fever lasting four weeks, accompanied by chills, myalgia, nausea, and jaundice in the last week. He was previously seen in a primary care center with antipyretics, without improvement. The physical examination showed a lucid patient with hypotension, tachycardia, and fever; epigastric and bilateral hypochondrial abdominal pain on superficial and deep palpation was evidenced. Hepatomegaly, jaundice, and petechiae were also found. Laboratory tests showed severe pancytopenia, acute renal failure, hydroelectrolytic disturbances, and hypoalbuminemia. The patient was admitted to the Pediatric Urgency service. After a poor progression, he passed away nineteen days after admission. The necropsy showed necrotic pseudomembranes in the colon and a histological image resembling the shape of a volcano, compatible with pseudomembranous colitis.

Evaluation of cell affinity on poly(L-lactide) and poly(epsilon-caprolactone) blends and on PLLA-b-PCL diblock copolymer surfaces.

An evaluation of cell proliferation and adhesion on biocompatible film supports was performed. A series of films were compression molded from commercially available poly (L-lactide), PLLA, and poly(epsilon-caprolactone), PCL, and from their melt mixed blends (PLLA/PCL blends). These were compared with compression molded films of PLLA-b-PCL model diblock copolymers. The samples were analyzed by differential scanning calorimetry (DSC), contact angle measurements, and scanning force microscopy (SFM). Cell adhesion and proliferation were performed with monkey derived fibroblasts (VERO) and with osteoblastic cells obtained either enzymatically or from explants cultures of Sprague-Dawley rat calvaria. Migration studies were performed with bone explants of the same origin. The results obtained indicate that although all materials tested were suitable for the support of cellular growth, a PLLA-b-PCL diblock copolymer sample with 93% PLLA was significantly more efficient. This sample exhibited a unique surface morphology with long range ordered domains (of the order of 2-3 mum) of edge-on PLLA lamellae that can promote "cell contact guidance." The influence of other factors such as chemical composition, degree of crystallinity, and surface roughness did not play a major role in determining cell preference toward a specific surface for the materials employed in this work.

Trypanocidal activity of abietane diterpenoids from the roots of Craniolaria annua.

A chloroform extract from roots of Craniolaria annua provided six new C-11 unsubstituted abietanes, 14-hydroxy-6,12-dione-7,9(11),13-abietatriene (1), 14-hydroxy-12-oxo-7,9(11),13-abietatriene (2), 6,12,14-trihydroxyabieta-5,8,11,13-tetraen-7-one (3), ar-abietatriene-12-ol-6,7-dione-14,16-oxide (4), ar-abietatriene-12,16-diol-14,16-oxide (5) and ar-abietatriene-12-ol-7-one-14,16-oxide (6), and two known compounds, ferruginol and stigmasterol. The structures of both the new and known compounds were established by spectroscopic methods. Abietane 1 gave 14-hydroxy-6-oxoferruginol (1A) upon treatment with NaBH4. Abietanes 1, 1A, 3-5 and ferruginol showed cytotoxic effects against trypomastigote and epimastigote forms of Trypanosoma cruzi and against fibroblastic Vero cells.

Serodiagnosis of chronic and acute Chagas' disease with Trypanosoma cruzi recombinant proteins: results of a collaborative study in six Latin American countries.

An enzyme-linked immunosorbent assay to diagnose Chagas' disease by a serological test was performed with Trypanosoma cruzi recombinant antigens (JL8, MAP, and TcPo). High sensitivity (99.4%) and specificity (99.3%) were obtained when JL8 was combined with MAP (JM) and tested with 150 serum samples from chagasic and 142 nonchagasic individuals. Moreover, JM also diagnosed 84.2% of patients in the acute phase of T. cruzi infection.

Trypanosoma cruzi ubiquitin as an antigen in the differential diagnosis of Chagas disease and leishmaniasis.

In the present report we describe Trypanosoma cruzi ubiquitin as an antigen to be utilized in the differential diagnosis of Chagas disease and leishmaniasis. Initially, recombinant T. cruzi ubiquitin was evaluated against a panel of sera by phage dot immunoassay, showing a good performance against chagasic sera. However, the presence of a carboxy-terminal tail region encoding a ribosomal protein homologous to a related protein present in the genome of Leishmania sp. gave significant cross-reactivity with leishmanial sera. Therefore, ubiquitin was purified by a simple biochemical protocol and its immunoreactivity was studied by enzyme-linked immunosorbent assay. Analysis of 104 sera indicates that the response to ubiquitin is very sensitive towards chronic chagasic sera (98%) and, more important, highly species-specific, presenting better performance compared to the use of the recombinant protein or the total epimastigote extracts when tested against a panel of leishmanial sera, where out of a total of 70 sera tested, only five sera from the mucocutaneous form of the disease reacted with T. cruzi ubiquitin. On the other hand, Leishmania ubiquitin was not recognized by chagasic sera, but was recognized by sera from different forms of leishmaniasis. These results make ubiquitin an excellent candidate to be used in the differential diagnosis of these two parasitic diseases. The molecular basis for this highly species-specific response is discussed.

Chromatin of Trypanosoma cruzi: in situ analysis revealed its unusual structure and nuclear organization.

Chromatin of Trypanosoma cruzi is known to be organized in classical nucleosomal filaments, but surprisingly, these filaments do not fold in visible chromosomes and the nuclear envelope is preserved during cell division. Our hypothesis about the role of chromatin structure in regulating gene expression and, more generally, cell functioning, pressed us to verify if chromatin organization is modulated during the parasite life-cycle. To this end, we analyzed in situ the fine structural organization of T. cruzi chromatin by means of an integrated biophysical approach, using differential scanning calorimetry and fluorescence microscopy. We observed that logarithmic forms exhibit a less condensed chromatin with respect to the stationary ones. Thermal analysis revealed that parasite chromatin is organized in three main levels of condensation, barring from the polynucleosomal filament till to superstructured fibers. Besides, the fluorescence images of nuclei showed a characteristic chromatin distribution, with defined domains localized near to the nuclear envelope. While in stationary parasites, these regions are highly condensed, in logarithmic forms they unfold by extending themselves toward the center of nucleus. These observations suggest that, in comparison with higher eukaryotes, in T. cruzi the nuclear envelope plays an unusual and pivotal role in interphase and in mitosis.

Differences in the nuclear chromatin among various stages of the life cycle of Trypanosoma cruzi.

Trypanosoma cruzi is the etiological agent of Chagas. Although the nuclear chromatin of this parasite is organized in the form of nucleosome filaments, its chromatin is physically and enzymatically fragile, and no condensation into chromosomes occurs during mitosis. All previous investigations have been carried out with epimastigote form in its proliferate stage. It is not known whether these differences in chromatin structure are also found in the non-proliferate stationary epimastigote forms and in tissue derived trypomastigotes. Our results confirm that chromatin of logarithmic epimastigotes presents limited compaction when increasing salt concentrations from 1 to 100 mM NaCl, and no 30-nm fibers were formed. Contrary to these results, non-proliferative forms of the parasites showed a pattern of compactation similar to that observed in rat liver chromatin, where solenoids of 30-nm fibers are formed at 100-mM NaCl. In accordance with these results, digestion of the nuclear chromatin with DNase I revealed that the chromatin of logarithmic phase epimastigotes was more accessible to the enzyme. We conclude from these results that structural differences in the chromatin exist not only between T. cruzi and higher eukaryotes but also among various forms of the parasite. The functional significance of these differences are currently under investigation.

Mechanisms of protein degradation in Trypanosoma cruzi

Proteolysis of endogenous proteins may play a key role in the adaptation of T. cruzi to the different host environments to which it is exposed during its complex life cycle. For this reason, we have attempted to study the intracellular pathways of protein degradation in the non infective epimastigotes form (EP strain) of T. cruzi. Following intracellular proteolysis by pulse chase experiments with 35 S methionine, we observed a significant inhibition (50 percent) of the degradation of endogenous proteins in log phase parasites in the presence of inhibitors of lysosomal functions, such as chloroquine and E 64. A significant increase in proteolysis was observed in stationary phase parasites which was reverted to log phase values by supplementing the chase medium with 0.5 percent glucose or 10 percent serum, or in the presence of chloroquine. Under this condition of nutritional stress, we could observe an increase in the activity of acid proteases. A significant increase in the degradation rates was observed when abnormal proteins were induced in the parasite by amino acid analogs and puromycin. This increase was not affected by E 64, suggesting the participation of non lysosomal mechanisms in the degradation of rapidly degradable abnormal proteins. Under these conditions, we could observe an increase in high molecular weight conjugates of ubiquitin with respect to endogenous proteins. These results suggest the importance of lysosomal mechanisms in the degradation of cellular proteins in nutritional optimal conditions and during nutritional deprivation, and the possible involvement of the ubiquitin system in the degradation of high turnover proteins