Pancreatic enzymes in serum and urine as indicators of pancreatic allograft rejection in the pig.

We have studied the reliability of serum and urinary immunoreactive anionic trypsin (irAT), immunoreactive cationic trypsin (irCT), and amylase activity as rejection indicators in a porcine whole-organ pancreaticoduodenal transplantation model with exocrine drainage to the urinary bladder. No immunosuppressive therapy was administered. Exocrine tissue integrity and function were studied by measuring these enzymes in serum and urine. Urine analyses were performed before and after an intravenous secretin-cholecystokinin stimulation. Of 16 transplanted pigs, 10 became diabetic during a 2-week observation period while six remained normoglycemic. Serum irAT was found to predict rejection while serum amylase and serum irCT did not. An increase in irAT was seen in rejecting pigs preceding the onset of hyperglycemia by a median of 2 days (range 1-9). Secretion of irAT into the urine remained high during the observation period in nondiabetic pigs while the output declined in diabetic pigs. This decline was seen after the increase in serum irAT. When urine was sampled after a secretin-CCK stimulation, these findings were clearly evident, but less unequivocal results were obtained without stimulation. IrAT measurements were superior to measurements of amylase, irCT, or bicarbonate. Thus rejection of a pancreatic allograft was first indicated by a temporary rise in serum immunoreactive anionic trypsin, probably due to the onset of tissue damage. Thereafter, stimulated urinary enzyme output levels gradually declined and finally, hyperglycemia developed.

Influence of prostanoids on gastrointestinal mucosal injury in experimental septic shock.

Capillary stasis and mucosal injury in the stomach and small intestine were studied in septic shocked pigs. Septicemia was induced by live E. coli i.v. in 28 animals. Additionally, five animals were infused with Ringer's solution and served as sham controls. The 28 E. coli-infused animals were pretreated with either a cyclooxygenase inhibitor--indomethacin, n = 6, a thromboxane (TxA2)-synthetase inhibitor--UK 38,485 alone, n = 6, or combined with a serotonin-antagonist--ketanserin, n = 9. Seven E. coli-infused animals were left untreated and served as septic controls. The sham controls were hemodynamically stable and had normal histological findings. All bacteria-infused animals exhibited signs of septic shock with pronounced hemodynamic reactions. Attenuation of the bacteria-induced increase in pulmonary arterial blood pressure was found in all pretreated animals but most pronounced in the indomethacin-pretreated group which also showed protection against gastric mucosal injury and capillary stasis. TxA2-inhibited animals had aggravated capillary stasis and mucosal injuries. It is concluded that gastric mucosal damage could be modified by drugs influencing the prostanoid system. The "cytoprotective" effect of prostaglandins seem to be of minor importance for the prevention of the gastro-intestinal mucosal injury seen in some series.

A comparison between IEEC, a new biodegradable particulate contrast medium, and iohexol in a tumor model of computed tomography imaging of the liver.

RATIONALE AND OBJECTIVES: Higher contrast between normal and pathologic tissues in the liver may enable detection of smaller lesions in computed tomography (CT). This can be obtained using a liver-specific contrast medium. The authors evaluate a new agent, IEEC (1'-Ethyloxycarbonyloxy)-ethyl-5-acetylamino-3-(N-methyl-acetylami no)-2,4,6- triiodo-benzenecarboxylate), in an animal model, as a potential contrast agent for CT scanning of the liver. The IEEC particulate contrast medium used is based on a prodrug ester design of metrizoic acid and accumulates rapidly in the liver. The particles are quickly degraded into well-known metabolites and excreted from the body. METHODS: Two groups of rabbits were inoculated with VX2-carcinoma directly into the liver by laparotomy. Computed tomography imaging studies were carried out 9 and 11 days after the inoculation. The investigation was designed as a crossover study. The first group was imaged both as controls (without contrast medium) and with the particulate contrast medium on the 9th day and with iohexol on the 11th day. The second group was imaged with iohexol on the 9th day and as controls, and with the particulate contrast medium on the 11th day. The contrast medium was administered in a dose of 100 mgI/kg. Iohexol was administered in a dose of 570 mgI/kg according to a standard clinical scheme in use at a radiology department for dynamic CT. Changes in normal liver/lesion contrast and the conspicuity of tumors were assessed. On completion of imaging studies on day 11, all animals were killed. The liver was removed and evaluated for the presence of tumors. RESULTS: At macroscopic inspection, all rabbits were found to have tumors ranging from 2 to 14 mm in diameter. The size and location of the tumors corresponded well with the CT images. In the images where the particulate contrast medium was used, the attenuation in the normal liver parenchyma and the contrast between normal liver and lesion was significantly higher compared with the images where iohexol was used or the controls. For all tumor sizes, the lesion detection capability with the particulate contrast medium was significantly higher compared with iohexol (P < .005) and controls (P < .05). CONCLUSIONS: VX2-carcinoma in rabbit liver is a useful model for studying the efficacy of contrast media in CT imaging. The particulate contrast medium IEEC improved visualization of liver tumors.

Heterogeneity of islet pathology in two infants with recent onset diabetes mellitus.

The mechanisms by which the beta cells of pancreatic islets are destroyed in insulin-dependent diabetes mellitus (IDDM) are poorly understood. In this report the pancreatic histo- and immunopathology of two children, both HLA-DR 3/4, DQ 2/8 positive and who both died from cerebral oedema within a day of clinical diagnosis of IDDM, were investigated. Patient 1, a 14-month-old girl, had a 4-week history of polydipsia and polyuria. Patient 2, a 3-year-old boy, had 2 days of illness. Both patients had a similarly severe loss of insulin cells but differed markedly as to the extent of lymphocytic islet infiltration (insulitis). Apart from insulitis, marked islet macrophage infiltration was demonstrated in both patients with the HAM-56 monoclonal antibody. Neither patient showed aberrant expression of HLA class II antigens on insulin-immunoreactive cells, but allele-specific HLA-DQ8 expression was evident on endothelial cells. Glutamic acid decarboxylase immunoreactivity was detected in both insulin- and glucagon-immunoreactive cells. It is concluded that the heterogeneity of islet pathology, especially insulitis, may reflect different dynamics and extent rather than different pathomechanisms of immune destruction of islets in IDDM.

The sequence of development of intestinal tissue injury after strangulation ischemia and reperfusion.

Tissue injury at reperfusion has been reported after partial ischemia. However, previous attempts to demonstrate a component of injury caused by reperfusion after total ischemia have failed. This study was performed to evaluate the hypothesis that in such situations the extent of the tissue injury caused by ischemia itself prevented detection of a reperfusion component. Rats were subjected to near-total intestinal ischemia by means of a hydrostatic pressure clamp that produced preferential venous occlusion (strangulation) for periods from 1 to 90 minutes. Tissue injury was evaluated microscopically by a blinded examiner. Ischemic periods of 20 minutes or less did not induce detectable tissue injury. Longer durations of ischemia caused villous injury: the longer the period of ischemia, the more extensive the tissue injury. However, there was no exacerbation of injury seen after reperfusion, regardless of the duration of ischemia. In a separate series of rats, total arterial occlusion was employed without concomitant venous congestion. Such isolation arterial occlusion of 40 to 60 minutes' duration was followed by a statistically significant exacerbation of tissue injury at reperfusion. Thus total intestinal ischemia may be followed by reperfusion injury if there is no concomitant congestion and if ischemic injury is not too extensive.

Islet cell antibody reactivity with human fetal pancreatic islets.

To evaluate the possibility of autoimmune processes against pancreatic islets in fetal life, we tested islet cell antibody (ICA) reactivity with 14 fetal pancreata obtained after abortion at the 15th up to the 19th week of gestation. Pancreatic islets positive for a monoclonal proinsulin antibody but non-reactive with ICA negative control serum were found in 9/14 pancreata and all (9/9) of them showed a positive reaction with the ICA standard. It is concluded that ICA reactivity may be detected in fetal human pancreata. Further studies on fetal islet cell antibody reactivity in the development of insulin dependent diabetes mellitus (IDDM) are warranted.

The course of neuroendocrine differentiation in prostatic carcinomas. An immunohistochemical study testing chromogranin A as an "endocrine marker".

To gain further insight into the pathogenetic aspects of neuroendocrine (NE) differentiation in prostatic carcinoma, the incidence of NE manifestations was studied during tumour progression in the course of the disease. This follow-up took the form of semiquantitative assessment of the NE cells in carcinomas by means of repeat biopsies at intervals of a few years, correlating the findings with those of conventional histopathological grading of the prostatic tumours. Immunoreactivity to chromogranin A (ChrA) and the Grimelius silver-staining technique were used to detect NE cells. A strong correlation was observed in all the 25 carcinomas studied between the results obtained with the Gimelius silver-staining and those obtained on the basis of immunoreactivity to ChrA. In addition, cells immunoreactive to an antiserum against ChrA found in virtually all sections from 24 cases of hyperplastic prostatic glands were found to be almost invariably argyrophil. Most of the 25 carcinomas underwent marked tumour progression, while the number of NE cells concomitantly increased. An unequivocal relationship can be stated between the degree of NE differentiation and tumour progression in our series of prostatic carcinomas treated with steroids-i.e., the more anaplastic the prostatic carcinoma, the more numerous are its NE cells. ChrA may be considered to be a sensitive marker for NE cells both in hyperplasia and in prostatic carcinomas.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunohistochemical investigations of beta-endorphin in human pancreatic islets.

The PAP technique was used to examine adult human pancreata (corpus) immunohistochemically for the presence of beta-endorphin containing cells. These cells were found to account for 4.8% of the islet cells. They are irregularly distributed within the islets, where they occur singly or in groups of 3 to 5 cells between the acini (0.4% of the parenchyma). Investigations designed to detect the simultaneous presence of beta-endorphin and somatostatin or glucagon revealed that beta-endorphin occurs in somatostatin cells (1.0% of the islet parenchyma). This is the 1st proof that these 2 hormones appear together. The simultaneous presence of beta-endorphin and glucagon in the same cell was also observed in 0.9% of the islet parenchyma. Earlier studies undertaken by us have shown that beta-endorphin is synthetized in the islets of Langerhans. Possible functions of beta-endorphin in the islets are discussed.

Beta-endorphin-immunoreactive cells in the human fetal pancreas.

This investigation has been carried out on 50 samples of fetal pancreata from the 10th to the 32nd week of gestation using the PAP technique. beta-Endorphin-reactive cells were morphometrically recorded by means of the point-counting method. beta-Endorphin reactivity occurred for the first time during the 15th week. During further development, beta-endorphin cells were found inside and outside the islets. From the 18th to 23rd week, these cells were primarily localized in the islet periphery. From the 24th week, they rearranged and occurred in irregular positions mixed with other islet cells. This rearrangement took place with a 4 week delay compared with the basic cell types of the islet organ. The extrainsular portion of these cells in the exocrine parenchyma varied between 0.3% in the 27th week and up to 10% in the 22nd week. Concerning the adult human pancreas, it has been suggested whether beta-endorphin cells may be a 6th basic cell type of the islet organ. Previous studies on the coexistence of somatostatin, glucagon and beta-endorphin in the same islet cell and the morphometric analysis would support this assumption. Biochemical examinations indicate that beta-endorphin is a modulator of insulin, glucagon, and somatostatin secretion in the islet organ. This is supported by the fact that beta-endorphin cells have extended cell bodies which is typical of cells with paracrine function.

Identification of monoclonal antibodies to pancreatic islet cells by immunoperoxidase staining.

Immunoperoxidase staining and enzyme-linked immunosorbent assay (ELISA) were used to identify monoclonal antibodies that reacted with pancreatic islet cells. All monoclonal antibodies produced against isolated human or rat pancreatic islets including one mouse autoantibody reacted with pancreatic islets in formalin-fixed pancreas sections, but not with rat kidney or thyroid. Reactivity was also found with suspensions of normal rat islet cells and rat insulinoma cells using a 3-stage immunoperoxidase procedure and an ELISA technique. Differences were observed in staining intensity between the various antigenic substrates tested suggesting variable cross-reactivity and/or number of epitopes. The sensitivity of the immunoperoxidase technique proved to be favourable for identification of monoclonal antibodies that recognize cellular constituents such as islet cell antigens present in low concentrations.

Neuron-specific enolase (NSE) as a neuroendocrine cell marker in the human fetal pancreas.

Using the PAP technique, we investigated the presence of neuron-specific enolase in the human fetal pancreas of 10, 12, and 14 weeks of gestational age. Neuron-specific enolase is present in the islet cells in the 10th week. Positive cells are situated mainly in duct epithelium. The number of cells with a positive reaction increases from the 12th to the 14th week. In the 14th week, they are clustered either near the ducts or between the acini. The numbers and localizations of the cells correspond to those obtained in previous studies with 4 basic islet cell types in the same material. The present results are a further proof that islet cells are biologically active during early fetal development.

Detection of proinsulin, C-peptide, insulin-A-chain, and glicentin in pancreatic islet cells of early human fetogenesis.

The presence of C-peptide, proinsulin, insulin-A-chain, and glicentin in human fetal pancreatic cells by using the PAP-technique was investigated and the results obtained compared with the occurrence of insulin or glucagon immunoreactive cells. In pancreatic sections obtained from 10 weeks old human fetuses we could identify cells reacting with antibodies directed against C-peptide, proinsulin, and insulin-A-chain. The majority of the cells were found in the duct epithelium and their number increased from the 10th to 14th week forming clusters near the ducts. The number and localization of the cells correspond exactly to the insulin positive cells. The presence of proinsulin and insulin-A-chains is a further proof of biological activity already in an early step of fetal development. The presence of glicentin-positive cells in the 10th week of gestational age as well as cells reacting with glucagon antibodies provide evidence for active glucagon biosynthesis. The number of these cells increased markedly in the 14th week of gestational age.

Argyrophil and beta-endorphin immunoreactive cells in focal islet-cell adenomatosis and insulin-producing islet-cell adenomata.

Pancreatic tissue from 3 cases of hyperinsulinemic hypoglycemia was examined using histochemical and immunoperoxidase staining techniques. The insular lesions present were adenomatosis and insulin-producing islet-cell adenomata. The great majority of the islet parenchymal cells in these lesions were reactive with antibodies to pro-insulin, C-peptide, and insulin. A variable number of islet cells was found to react with beta-endorphin antiserum in all 3 cases, while the reaction with antiserum against the neural tissue marker antigen, S-100, was restricted to the cases with islet-cell adenoma. Argyrophil parenchymal cells were present in focal adenomatosis but almost absent in insulomata. These results suggest that various lesions of the endocrine pancreas causing hypoglycemia can be distinguished by means of specific histo- and immunocytochemical methods because of differences in the distribution of characteristic cellular antigens.

Primary intracranial epidermoid carcinoma.

The cytologic findings in the cerebrospinal fluid in a case of primary intracranial epidermoid carcinoma are presented. The presence of a few squamous cells showing hyaline cytoplasm compatible with keratinization suggested the possible nature of the tomographically observed lesion. These tumors may arise in congenital epidermoid cysts at the base of the brain. Because of their location and meningeal spread, such tumors may cause severe symptoms and death despite a minute size, as illustrated by this case.

A simple method for processing cytologic samples obtained from body cavity fluids and by fine needle aspiration biopsy for ultrastructural studies.

A simple method for processing routine cytologic samples for electron microscopy is described. The method has been successfully applied to body cavity fluids and fine needle aspiration biopsies. The technique neither requires additional time and personnel nor interferes with the normal activity of the laboratory.

Ontogeny of the pancreatic islet parenchymal cells in the rabbit--an immunohistochemical and ultrastructural study with particular regard to the earliest appearance of argyrophil insulin-immunoreactive cells.

Six early developmental stages of the rabbit pancreas were selected, viz. the embryonic ages 10 days 10 hours, 10 days 18 hours, 11 days 14 hours, 13 days, 15 days, and 18 days. Both non-immunological (histologic-tinctorial features, including argyrophilia, and transmission electron microscopy) and immunohistochemical (the indirect immunofluorescence and/or the peroxidase-anti-peroxidase procedure) methods were used to follow the time-course for the appearance and differentiation of both endocrine (islet) cells and exocrine acinar epithelium. The immunological procedures were, however, limited to the 3 later developmental stages. In the first 3 developmental stages only the dorsal anlage of the pancreas could be found. It was just investigated ultrastructurally. Then, a few parenchymal cells were observed, equipped with secretory granules of endocrine type, indicating that an early differentiation of islet cells had already begun. In the later 3 developmental stages a ventral pancreas anlage was present and at least 2 types of argyrophil islet cells, equipped with secretory granules, were observed. In the pancreas anlage of 13-day-old embryos these early endocrine cells were found to be glucagon-immunoreactive. At the developmental age of 15 days argyrophil insulin-immunoreactive cells were also present, and in the 18-day-old embryos a few somatostatin cells could occasionally be discovered, too. No PP cells were found. Any exocrine acinar differentiation (with zymogen granules) was not observed until at the developmental age of 18 days.

Effects of a calcium antagonist (nifedipine) on cats in live E. coli bacteriemic shock.

The effects of a calcium antagonist, nifedipine, on cardiovascular reactions and on gastrointestinal mucosal integrity was studied in a standardized feline bacteriemic model. Nifedipine pretreatment delayed the development of cardiovascular derangement and reduced the severity of the intestinal but not the gastric mucosal injury. The effect on the intestinal mucosa could be due to the delayed development of hypotensive shock but also to a protective effect on the superficial mucosal cells.