Normal Dog Pancreas Research Articles

Metallic Stent Placement in the Pancreatic Duct: An Experimental Study in the Normal Dog Pancreas

To evaluate radiologic and histologic changes in the pancreatic duct and parenchyma after placement of metallic stents in normal dog pancreatic ducts. Eight dogs underwent duodenotomy and placement of a balloon-expandable metallic stent 1.5 mm in diameter and 1 cm in length in the main pancreatic duct. Two dogs each were killed at 1 day and 1, 3, and 5 months after stent placement. Pancreatic duct injection images were obtained before and immediately after stent placement and just before and after sacrifice. Macroscopic and microscopic examinations were also performed. Pancreatic duct injection images revealed mild dilation of pancreatic ducts related to the procedure immediately and 1 day after stent placement. Microscopically, denudation of the ductal epithelium and small foci of fat necrosis were observed. At 1 month or later, ductal stricture (n = 4) and occlusion (n = 2) were found at the downstream edge of the initially stent-implanted region associated with dilation of the duct upstream. Epithelial hyperplasia was observed at the sites of ductal stricture and occlusion at 1 and 3 months. Stents remained in the region in which they were initially placed until 1 month. However, stents had moved downstream from their initial positions at 3 and 5 months. One stent had migrated into the duodenum at 5 months. Stone formation was found in the ductal lumen and around the stents at 3 and 5 months. Ducts and parenchyma became fibrotic proportional to the period of stent implantation. Metallic stent placement in the pancreatic duct causes ductal and parenchymal damage related to the procedure and reaction to the stent in the normal dog pancreas.

Polyethylene pancreatic duct stent-induced changes in the normal dog pancreas

This study was undertaken to evaluate the frequency, severity, and reversibility of stent-induced pancreatic ductal and parenchymal changes in the normal dog pancreas. Six adult mongrel dogs underwent duodenotomy and placement of 5F polyethylene pancreatic duct stents into the main pancreatic duct. After 8 weeks of stenting, the animals were randomly assigned to one of three groups: group I (n = 2), sacrifice; group II (n = 2), stent removal followed by sacrifice after an 8-week recovery period; group III (n = 2), the stent was exchanged and kept in place for an additional 8 weeks. In group III at 16 weeks the stent was removed, and the animals were allowed to recover for 8 weeks before sacrifice. Pancreatograms were obtained at each operation and were normal before stent placement. Gross and histologic evaluation was performed at the time of sacrifice. All stented animals developed radiographic, gross, and histologic abnormalities. Pancreatograms showed duct dilation in the stented region associated with a short stenosis at the mid to upstream segment of the stent. Group III animals had more advanced radiographic changes than group I and II animals. The radiographic findings were associated with gross evidence of fibrosis, which increased proportionately with the length of the stenting period. Group III animals developed moderate to severe pancreas atrophy. Histologic changes of obstructive pancreatitis were present in most experimental dogs. The results of this study suggest that pancreatic stenting may cause permanent damage in the normal dog pancreas. The cause of the damage appears to be related to stent occlusion, perhaps exacerbated by local stent-induced trauma.

Plasma responses of pancreatic polypeptide, glucagon and insulin in normal and alloxan diabetic dogs, and their regional levels in the pancreas

The plasma responses of pancreatic polypeptide (PP), glucagon (IRG) and insulin (IRI) after administration of beef soup were studied in normal and alloxan diabetic dogs kept in a poor metabolic state for 4 weeks, and their regional levels in the pancreas were determined and compared at the uncinate process, head, body and tail. The plasma PP levels of the diabetic dogs were significantly higher than those of the normal dogs and they increased after beef soup administration in both groups. The plasma IRG levels did not change significantly after beef soup administration in either group, but significantly high levels were found in the diabetic dogs. The IRI content in the normal dog pancreas was highest at the tail, followed in order by the body, head and uncinate process, and was decreased to about one-tenth to one-fortieth in the alloxan diabetic dogs. On the other hand, the amounts of PP in the pancreas of both normal and diabetic dogs were greatest at the uncinate process, followed in order by the head, body and tail. In an inverse relationship with the PP findings, the IRG content in the pancreas was highest at the tail and lowest at the uncinate process in both kinds of dogs. The differences in both PP and IRG between diabetic and normal dogs were generally not significant. These results show that irrespective of the high plasma levels of PP and IRG in the diabetic dogs, their levels in the pancreas did not change significantly.

Secretin inhibits glucagon in the isolated perfused dog pancreas.

To investigate the effect of secretin upon islet hormone secretion, highly purified porcine secretin was perfused at 0.0001, 0.001, 0.01, and 0.1 clinical unit (CU)/ml in the isolated normal dog pancreas. The lowest concentration of secretin, a level which is within the physiologic range, significantly suppressed glucagon release and stimulated somatostatin release but was without significant effect on insulin release. At each concentration of secretion tested glucagon was dramatically suppressed and somatostatin was significantly stimulated, both in a dose-dependent manner. B-cell activity was first augmented by the concentration of 0.001 CU/ml (94 pM) of secretin. These data demonstrate that physiologic levels of secretin suppress pancreatic A-cell activity in the isolated dog pancreas.

Stimulatory effect of beta-hydroxybutyrate on the release of somatostatin from the isolated pancreas of normal and streptozotocin-diabetic dogs.

The present investigation was undertaken to ascertain whether the ketone body, beta-hydroxybutyrate (BOH), affects the somatostatin secretion from the isolated pancreas of normal and streptozotocin (STZ)-diabetic dogs. We found (1) that the addition of 10 mM DL-BOH to the perfusate augmented somatostatin secretion from the normal dog pancreas. The responses obtained were, however, modified by the prevailing glucose level with higher responses appearing at high (11 mM) rather than at zero glucose. The effect of BOH could not be attributed to changes in osmolarity, Na+, or H+ concentrations. (2) BOH at doses well within the physiologic and pathophysiologic range stimulated the secretion of somatostatin from the normal pancreas in a dose-dependent manner. The threshold concentration was close to 1 mM corresponding to 0.5 mM of the biologically active D form. (3) In the STZ-diabetic dogs, BOH (1-20 mM) elicits a dose-related enhancement of the pancreatic somatostatin secretion. In the two groups, identical D-cell responses were obtained at 3, 10, and 20 mM BOH. The results demonstrate that physiologic levels of BOH stimulates the release of somatostatin from pancreas of normal and STZ-diabetic dogs, thereby providing support for a physiologic influence of BOH over the secretory function of the pancreatic D-cell. The possible existence of a BOH-somatostatin feed-back mechanism operating in the diabetic organism is discussed.

Preparation of viable islet cells from dogs by a new method.

A new method for the preparation of viable islet cells from the normal dog pancreas is described, based on the perfusion of the pancreatic duct with collagenase. Exposure of the acinar tissue to the highest concentrations of collagenase results in improved islet yield with decreased acinar contamination. After the selective digestion of the gland by ductal perfusion, mechanical dissociation liberates single cells and clumps. Analysis of the procedure by insulin yield and amylase attrition indicates a 57% B-cell recovery and a sixfold enrichment in B-cell concentration. The cells have been transplanted as autografts into dogs after pancreatectomy. In 5 of 7 transplanted dogs, normoglycemia was achieved postoperatively.

Somatostatin-like polypeptides in the plasma and tissues of a patient with a somatostatin-producing tumour.

The existence of multiple molecular forms of the polypeptide hormones and probably of all secretory proteins is a consequence of their mode of biosynthesis. Polypeptides larger than somatostatin (mol.wt. 1600) that possess somatostatin-like immunoreactivity have been detected in extracts of hypothalamus, extrahypothalamic brain, stomach, pancreas and gut. Analysis of newly synthesized proteins in pancreatic islets of the anglerfish (Noe et al., 1979) and rat (Patzelt et al., 1979) has indicated a mol.wt. of approx. 12000 for prosomatostatin. Release into the circulation of multiple forms of a hormone is a characteristic of endocrine neoplasms and has been extensively documented in the case of islet tumours. The clinical and morphological features of a patient with a somatostatinproducing tumour of the pancreas and multiple liver metastases has recently been described (Krejs et al., 1979). In the present study, the distribution of molecular forms of the somatostatin-like polypeptides in extracts of the primary tumour of this patient are compared with those released by the primary tumour and by the metastases into plasma. Blood samples, collected from a peripheral vein by the method of Harris et al. (1978), were taken from the patient in the fasted state and after a 4.2MJ (1000 calorie) mixed meal. Further samples were taken, in the fasted state, after resection of the primary tumour. Tissue was extracted by the method of Conlon et al. (1978a). Plasma samples and tissue extracts were chromatographed on columns of Biogel P-10 and Sephadex G-200 and somatostatin-like polypeptides in the efRuent fractions were detected by radioimmunoassay (Harris et al., 1978). Samples were diluted at least 20-fold before assay so that degradation of the radio-labelled tracer and non-specific interference by plasma proteins in the assay were negligible. Amounts of immunoreactivity equivalent to 9-25 ng of somatostatin/ml were measured in plasma samples in the fasted state (value in normal subjects < 100pg/ml) and increased 8-fold after a mixed meal. Three samples of tissue from the primary tumour contained immunoreactivity equivalent to 0.8, 1.2 and 4.6pg of somatostatin/mg wet wt. of tissue (the normal pancreas content was 3 1440pg/mg wet wt. of tissue; McIntosh et al., 1978). The somatostatin-like polypeptides in plasma samples and tissue extracts were resolved by gel filtration into three main peaks in the 10000-15000, 250Ck3500 and 1500-2000 mol.wt. zones. The relative amounts of these components were estimated from the area under the peaks and are shown in Table 1. Plasma samples, but not tissue extracts, contained low amounts (< 1% of total immunoreactivity) of somatostatin-like immunoreactive material, which was eluted from Sephadex G-200 columns in the 150000-200000 mol.wt. zone and may represent smaller immunoreactive peptides bound non-covalently to plasma proteins (Conlon et al., 1978b). The amounts of the higher molecular-weight components relative to the somatostatin-sized component are higher in plasma than in tissue extracts, which probably reflects a lower rate of clearance of the larger peptides from the circulation. The neoplasms responded to a nutrient stimulus with an increased release of all three components, but the rate of release of the 1500-2000-mol.wt. component was much greater than that of the larger peptides. The marked increase in the relative amounts of the 1000015 000and 2500-3500-mol.wt. polypeptides in the postoperative plasma is evidence for an increased rate of release of these components from the metastases compared with the primary tumour. Somatostatin-like polypeptides of mol.wts. approx. 12000, 3000 and 1600 have been identified in extracts of normal dog pancreas (Conlon et al., 1978a), but only the 1600-mol.wt. component was detected in dog plasma and in the plasma-free efAuent of the isolated dog pancreas (Conlon et al., 19786). Proteolytic cleavage at sites of pairs of basic amino acids represents the predominant, but not exclusive, means of processing of prohormones. In the light of evidence from biosynthetic studies, it is proposed that the component with mol.wt. approx. 12000 in normal and tumour tissue represents prosomatostatin, which is specifically cleaved to the approx. 1600-mol.wt. component. The approx. 3000-mol.wt. polypeptide may be a product of non-specific cleavage of the prohormone. In normal pancreatic tissue, the rate of processing of prosomatostatin is rapid so that concentrations of the prohormone and the approx. 3000-mol.wt. component are low and only the 1600-mol.wt. component is released into plasma. In the tumours, however, the rate of conversion of the prohormone into hormone is much slower, so that high concentrations of prosomatostatin and its non-specific cleavage products accumulate in the cells and are subsequently released into plasma. The rate of processing of prosomatostatin appears to be less in the metastases than in the primary tumour.

Pancreatic immunoreactive somatostatin release.

The location of the somatostatin-containing D-cells of the pancreatic islets between the A- and B-cells suggests that their function might be to inhibit insulin and/or glucagon secretion by these neighboring cells. To determine if insulin and/or glucagon, in concentrations that might be present in the extracellular space surrounding the D-cells, stimulate immunoreactive somatostatin (IRS) release, we perfused 10 microng of glucagon or 10 milliunits of insulin per ml in 11 isolated dog pancreases, for 40 min in seven experiments and for 100 min in four experiments. In eight of the nine experiments in which glucagon was perfused, a prompt and significant rise in mean IRS release, ranging from 71 to 128% above the control level, was observed. In the eight experiments in which insulin was perfused. IRS did not increase during the first 40 min; in the two 100-min insulin experiments, it did rise during the final 50 min, however. To determine the effect of an A- and B-cell secretogogue on IRS release, we perfused 20 mM arginine for 60 min in six experiments. In all, IRS rose within 3 min and reached a level 71-465% above the control, remaining significantly elevated throughout the perfusion, while glucagon and insulin rose to peak levels at 2 min and then declined somewhat despite continuing arginine perfusion. The results indicate that perfusion of the normal dog pancreas with high doses of glucagon or arginine is accompanied by a prompt increase in IRS release and are compatible with a local feedback circuit involving A- and D-cells. Insulin appears not to augment IRS release, at least not promptly, but IRS stimulated by local endogenous glucagon could inhibit the B-cell response to locally secreted glucagon and thereby influence the composition of the insulin/glucagon secretion mixture.

Microcirculation of the normal and inflamed canine pancreas.

Pancreas of normal dogs and the inflamed gland of experimental pancreatitis were studied by intra-arterial injection of Microfil, a silicone-rubber compound especially suited for study of the microcirculation. Duodenal vasculature and lobular vessels of the pancreas were studied as were those supplying the duct. Inter-lobular vessels were well visualized in the normal pancreas and intralobular vessels formed a fine reticular pattern throughout the cleared lobules. A complex network of vessels in the pancreatic duct was observed using this technique, apparently derived from the interlobular vessels. The blood supply of each layer of the duodenum was evaluated. Intraductal trypsin injection produced focal areas of pancreatitis associated with edema, poor vascular filling and spastic changes of the lobular vessels. Extravasation of Microfil, although not apparent on normal specimens, was prominent in inflamed specimens and suggested vascular weakness and disruption. Pancreatic lobules adjacent to inflammatory areas showed definite evidence of dilatation. The inflamed pancreatic ducts were markedly edematous, thickned, and showed incomplete vascular filling. When the duodenum adjacent to pancreatitis was injected with Microfil, edema and vasoconstriction were especially prominent in the duodenal muscular layer.

The dog pancreatic X cell: A light and electron microscopic study

AbstractA light and electron microscopic study was conducted on the X cell of the normal dog pancreas. These cells were identified by their acidophilic cytoplasmic granules which do not stain with iron or phosphotungstic acid hematoxylin and also show a distinctive multilobate nucleus. They were found to be neither argyrophilic nor argentaffinic. Ultrastructurally, when fixed in osmium alone, the X cell cytoplasm contained numerous rounded, smooth membrane bound secretory granules with content of low electron density. However, after double fixation in glutaraldehyde and osmium the granules appeared irregularly round, ovoid or kidney shaped with content which may vary from electron lucent to homogeneously electron dense with gradations between. The functional significance of the X cell is unknown. However, its structure showed it to be a distinct, active secretory element, independent of other pancreatic exocrine and endocrine cell types.

Corpuscles containing crystalline formations in the exocrine pancreas of the dog

IN studies pursued of the ultrastructure of experimental pancreatitis [3, 41, we were able to observe a significant increase in the number of intracytoplasmic crystalline formations in the acinar cells of the dog pancreas after intravenous injection of heterological serum. This observation bears on small pieces of 14 dog pancreas taken under general anaesthesia with Penthiobarbital of sodium, fixed in 2 per cent osmium tetroxide (phosphate buffer) embedded in Epon. The stain used was lead-hydroxide. Complex bodies were observed, measuring from 1 to 2 p in diameter, each separated from the surrounding cytoplasm by its own membrane having two or three layers. The formations themselves are of a heterogenous nature. They are composed of three types of elements (Fig. I): tridimensional crystals; highly osmiophilic, homogeneous and rounded droplets, probably of lipid nature; a heterogenous granular matrix. The crystals are made up of a uniform alignment of granules 110 A in average diameter separated from each other by a distance of approximately 160 A. The same cross section frequently shows crystals of different orientations (Fig. 2). What is the significance of these crystals? Their presence in the normal dog pancreas (7 animals) was observed in very rare cases. In contrast in 3 dogs, their number increased markedly in most cells after intravenous injection of 20 cc of rabbit serum containing anti-dog pancreas antibodies. Their multiplication has no relation, however, to a specific phenomenon of immunization, since the same results were obtained after intravenous injection, with the same dosage of previously non-immunized rabbit serum (1 animal), or after intravenous injection of previously non-immunized horse serum (1 animal). On the other hand, their appearance seems also to have no relation to a simple phenomenon of shock, since the injection of Histamine in sublethal doses (1 animal), as well as the intravenous injection of 20 cc of a solution of crystallized Ovalbumine diluted to 50/1000 (1 animal), does not increase their frequency. More over, Ichikawa [7] did not observe such crystals in the canine pancrease after hormonal stimulation. By morphological analogy with Tropomyosine crystals and those described as occurring at the time of ovogenesis of the Planorbis, these formations seem to us to be crystals of protein nature [2].

REGIONAL DIFFERENCES IN THE GLUCAGON CONTENT OF PANCREAS FROM ALLOXANIZED DOGS

The uncinate process of the dog pancreas is devoid of A cells. Pancreatic extracts from the uncinate process of alloxanized dogs showed neither hypernor hypo-glycemic activity. These findings suggest that the lack of hyperglycemic effect of extracts from the uncinate process of the normal dog pancreas (11) simply indicates absence of glucagon and is unrelated to the presence of insulin. These results are in agreement with the hypothesis that glucagon is originated by the A cells of the pancreas. Neither the D nor X cells showed any sign of damage following alloxan injection during the period of observation.

REGIONAL DIFFERENCES IN THE GLUCAGON CONTENT OF PANCREAS FROM ALLOXANIZED DOGS

The uncinate process of the dog pancreas is devoid of A cells. Pancreatic extracts from the uncinate process of alloxanized dogs showed neither hypernor hypo-glycemic activity. These findings suggest that the lack of hyperglycemic effect of extracts from the uncinate process of the normal dog pancreas (11) simply indicates absence of glucagon and is unrelated to the presence of insulin. These results are in agreement with the hypothesis that glucagon is originated by the A cells of the pancreas. Neither the D nor X cells showed any sign of damage following alloxan injection during the period of observation.