Structural basis for delta cell paracrine regulation in pancreatic islets.

Little is known about the role of islet delta cells in regulating blood glucose homeostasis in vivo. Delta cells are important paracrine regulators of beta cell and alpha cell secretory activity, however the structural basis underlying this regulation has yet to be determined. Most delta cells are elongated and have a well-defined cell soma and a filopodia-like structure. Using in vivo optogenetics and high-speed Ca2+ imaging, we show that these filopodia are dynamic structures that contain a secretory machinery, enabling the delta cell to reach a large number of beta cells within the islet. This provides for efficient regulation of beta cell activity and is modulated by endogenous IGF-1/VEGF-A signaling. In pre-diabetes, delta cells undergo morphological changes that may be a compensation to maintain paracrine regulation of the beta cell. Our data provides an integrated picture of how delta cells can modulate beta cell activity under physiological conditions.

Pancreatic islet of Langerhans' cytoarchitecture and ultrastructure in normal glucose tolerance and in type 2 diabetes mellitus.

While a number of structural and cellular abnormalities occur in the islet of Langerhans in diabetes, and in particular in type 2 diabetes, the focus has been mostly on the insulin producing ß-cells and only more recently on glucagon producing α- and δ-cells. There is ample evidence that in type 2 diabetes mellitus (T2DM), in addition to a progressive decline in ß-cell function and associated insulin resistance in a number of insulin-sensitive tissues, alterations in glucagon secretion are also present and may play an important role in the pathogenesis of hyperglycemia both in the fasting and in the postprandial state. Recently, a number of studies have showed that there are also functional and structural alterations in glucagon-producing α-cells and somatostatin-producing δ-cells. Thus, it is becoming increasingly clear that multiple cellular alterations of multiple cell types occur, which adds even more complexity to our understanding of the pathophysiology of this common and severe disease. We believe that persistent efforts to increase the understanding of the pathophysiology of hormone secretion in the islets of Langerhans will also improve our capability to better prevent and treat diabetes mellitus.

Beta-cell destruction and preservation in childhood and adult onset type 1 diabetes.

Previous studies describing the symptomatic onset of type 1 diabetes (T1D) and rate of beta-cell loss (C-peptide) support the notion that childhood onset T1D exhibits more severe beta-cell depletion compared to adult onset T1D. To test this notion, we performed whole pancreas analyses in two T1D cases, one of childhood onset (7-year old, onset at 1.5-year) along with an adult onset case (43-year old with onset at 27-year). Both cases were matched for age and gender with control subjects. Striking regional differences in beta-cell loss were observed in both T1D cases, with severity of loss in the order of tail > body > head regions. In contrast, pancreatic alpha- and delta-cell mass was similar in controls and T1D patients. In the childhood onset T1D case, no intra-islet beta-cells were detected while in the adult onset case, beta-cell containing islets were found, exclusively in the head region. In the latter case, considerable numbers of small cellular clusters negative for three major endocrine hormones were observed, in islets with or without beta-cells. Ultrastructural analysis suggests these cells correspond to degenerating beta-cells, with empty granular membranes and abnormal morphology of nuclei with intranuclear pseudo-inclusions, adjacent to healthy alpha- and delta-cells. These results support a hypothesis that during T1D development in childhood, beta-cells are more susceptible to autoimmune destruction or immune attack is more severe, while beta-cell death in the adult onset T1D may be more protracted and incomplete. In addition, T1D may be associated with the formation of "empty" beta-cells, an interesting population of cells that may represent a key facet to the disorder's pathogenesis.

Small human islets comprised of more ß-cells with higher insulin content than large islets.

For the past 30 years, data have suggested that unique islet populations exist, based on morphology and glucose sensitivity. Yet little has been done to determine the mechanism of these functional differences. The purpose of this study was to determine whether human islets were comprised functionally unique populations, and to elucidate a possible mechanism. Islets or pancreatic sections from 29 human donors were analyzed. Islets were isolated and measured for insulin secretion, cell composition and organization, insulin and glucagon granule density and insulin content. Insulin secretion was significantly greater in small compared with large islets. In sectioned human pancreata, ß-cells comprised a higher proportion of the total endocrine cells in small islets (63%) than large islets (39%). A higher percentage of ß-cells in small islets contacted blood vessels (44%) compared with large islets (31%). Total insulin content of isolated human islets was significantly greater in the small (1323 ± 512 µIU/IE) compared with large islets (126 ± 48 µIU/IE). There was less immunostaining for insulin in the large islets from human pancreatic sections, especially in the core of the islet, compared with small islets. The results suggest that differences in insulin secretion between large and small islets may be due to a higher percentage of ß-cells in small islets with more ß-cells in contact with blood vessels and a higher concentration of insulin/ß-cell in small islets.

Pancreatic D-cells in aging and intraislet effects of pancreatic somatostatin.

In old organisms pancreatic D-cells are not changed in number. During the aging in mentioned cells takes place the intensification of secretory and extrusive functions, which are more prominent in old organisms than in young ones. Peripherally situated D-cells are vascularly ineffective within the pancreatic islet and do not suppress locally B- and A-cells. D-cells' major target tissue may be pancreatic acinar cells. Functionally activated D-cells in old organisms may play the main role in the development of involutive processes in exocrine pancreas and in its atrophy. Stagnation of the secretory granules in pancreatic A- and B-cells in old ages could not be caused by influence of paracrine effect of somatostatin. The given process could be considered as a result of reduction of energopotentials and suppression of signal ways for initiation of insulin and glucagon secretion. Respectively, extrusion impediment of secretory granules resulted in their stagnation could be explained by suppression of exocytosis as an energy- and signal-dependent process. We suppose that cytotopographic and microvascular peculiarities of pancreatic islets in human beings and rodents is a reflection of intensification of insulin apparatus and is directed to loose the B-cells from the local (microvascular or paracrine) influences (effects of D- or A-cells). The mentioned is of high physiological importance (especially in the process of aging) for the organisms of above-presented taxonomic groups due to rich amount of carbohydrates in their food ration. The above-mentioned fact gains the special importance in human beings, where evolutionary "solitary" (represented by single B-cells) insulin apparatus is faced with evolutionary "rooted" strong and diverse contrainsulin apparatus, leading to development of diabetes mellitus (type 2) in late ages.

Intranuclear rodlets in human pancreatic islet cells.

OBJECTIVES: Intranuclear rodlets (INRs) are rod-shaped intranuclear inclusions that we have described in neurons of the human brain. We recently identified these structures in pancreatic islet cells. The objectives of this study are to describe the light microscopic features and cellular pattern of distribution of INRs in human pancreatic islet cells. METHODS: Double immunofluorescence staining was performed on 5 human pancreatic tissue samples for the detection of class III beta tubulin (C3T) to detect INRs and for promyelocytic leukemia (PML) protein to examine the relationship between PML and INRs. RESULTS: Intranuclear rodlets were detected in 22.99% of pancreatic B cells compared with only 3.11%, 1.80%, and 1.60% of A, D, and PP cells, respectively. Twenty-four percent of C3T-immunoreactive INRs showed partial or complete immunoreactivity for PML. Promyelocytic leukemia staining within the nuclei of B cells was confined to INRs and was not present in the typical PML bodies present in other cell types. Spatially, PML and C3T staining of islet cell INRs appeared to be mutually exclusive within individual INRs. CONCLUSIONS: Intranuclear rodlets are present within the nuclei of pancreatic islet cells, where they reside predominantly but not exclusively in B cells. Immunoreactivity of B-cell INRs for PML suggests that the functional significance of INRs may be related to that of PML and/or PML bodies. Conversely, the exclusive localization of PML staining to INRs in B cells indicates that PML's function in B cells is selectively associated with INRs. The mutually exclusive pattern of PML and C3T staining suggests dynamic interactions between these 2 proteins in B-cell INRs. In light of evidence for the involvement of INRs and of PML bodies in disease, it will be of interest to investigate these structures in animal models of diabetes and in human diabetes.

G and D cells in rat antral mucosa: an immunoelectron microscopic study.

AIM: To investigate the gastrin secreting cells (G cells) and the somatostatin secreting cells (D cells) of antral mucosa in rats at the ultrastructural level. METHODS: Revised immunoelectron microscopic technique was used to detect the G cells and D cells in rat antral mucosa through gastrin and somatostatin antibodies labeled by colloidal gold. Also the relevant quantitative analysis regarding the granular number of colloidal gold in G cells and in D cells was conducted. RESULTS: Immunological granules of colloidal gold were distributed in G cells and D cells. Gastrin labeled golden granules or somatostatin labeled ones presented mainly as lobation-like or island-like congeries. Most of the golden congeries were observed dissociated in cytoplasms of G cells or D cells, near the basement membrane. A few golden congeries were located in nuclei. The number of golden granules in one G cell was around 107.04 +/- 19.68 and was 83.36 +/- 17.58 in one D cell. CONCLUSION: Gastrin secreting granules are located in cytoplasms and nuclei of G cells, and somatostatin secreting granules both in cytoplasms and in nuclei of D cells. The number of golden granules can be quantitatively analyzed to determine the relative amount of gastrin secreting granules or somatostatin secreting granules.

Ultrastructural immunohistochemical localization of gastrin, somatostatin and serotonin in endocrine cells of human antral gastric mucosa.

Five types of endocrine cells are found in the human antral gastric mucosa: gastrin (G) cells, somatostatin (D) cells, enterochromaffin (EC) cells and cells with an unknown secretory product (D1 cells and P cells). The content of secretory granules, gastrin, somatostatin and serotonin, was evaluated using electron microscopic immunohistochemistry and was compared with the granular content in G cells, D cells and EC cells as determined by routine electron microscopy. Semi-quantitative scoring of the granular content was performed on a scale 1-4 (empty-full). The content of gastrin (2.5 +/- 0.2) and somatostatin (3.3 +/- 0.2) in the granules was not different from the granular content in G cells (2.5 +/- 0.3; p > 0.05) and D cells (3.5 +/- 0.2; p > 0.05). Gastrin was also found in G cells in a nongranular form. The content of serotonin in granules (2.8 +/- 0.3) was smaller than the granular content in EC cells (3.7 +/- 0.2; p < 0.05). In intermediate-full and intermediate-empty granules, serotonin was localized in the periphery of granules whereas the granular content in EC cells was localized in an eccentric or central pattern. The granular content of D1 cells and P cells was 3.8 +/- 0.2, and 3.4 +/- 0.2, respectively. It is concluded that gastrin and somatostatin immunostaining in granules of G cells and D cells reflects the granular content in G cells and D cells, respectively, whereas serotonin immunostaining does not agree with the granular content of EC cells.

Somatostatin and D cells in patients with gastritis in the course of Helicobacter pylori eradication: a six-month, follow-up study.

BACKGROUND/AIMS: As well as causing chronic gastritis, Helicobacter pylori predisposes patients to peptic ulcer disease and gastric cancer, and induces gastric functional disorders. The aim of our study was to investigate the effects of H. pylori eradication therapy on the morphological and functional recovery of gastric antral and corpus D cells in patients with chronic gastritis during 6 months of follow-up. PATIENTS AND METHODS: Forty consecutive, dyspeptic patients referred for endoscopy (31 with H. pylori infection and nine controls; mean age 49 years; 17 men, 23 women) entered the study. All patients had histological signs of gastritis but no signs of peptic ulcer or gastric cancer. Antrum (n=8) and corpus (n=6) biopsy specimens were collected for routine histology, radioimmunoassay tissue somatostatin levels, immunohistochemistry and electron microscopy, prior to and 6 months after therapy. Basal plasma somatostatin levels were determined prior to eradication, plus 6 weeks and 6 months after therapy. Eradication therapy consisted of amoxicillin, metronidazole and omeprazole. RESULTS: Basal somatostatin plasma values in antral and corpus tissue were lower in infected patients than in the H. pylori-negative controls at the beginning of the study. A significant increase occurred after successful eradication therapy, together with an increase in the number of D cells in both regions. Changes in the D-cell ultrastructure in antral and corpus mucosa after eradication therapy suggest an increase in somatostatin synthesis and secretion. CONCLUSIONS: The structural and functional restoration of D cells following eradication therapy indicates possible recovery of the diseased mucosa.

Electron microscopic immunohistochemical investigation of chromogranin A in endocrine cells in human oxyntic gastric mucosa.

Human oxyntic gastric mucosa harbours 6 types of endocrine cells: enterochromaffin-like (ECL) cells, enterochromaffin (EC) cells, somatostatin (D) cells, and cells with an unknown secretory product (P cells, D1 cells and X (A-like) cells). In the present study, intracellular localization and granular content of chromogranin A (CGA) in these cells have been investigated by electron microscopic immunohistochemistry. The content of CGA in granules of the various types of endocrine cells was evaluated and compared with the content of serotonin and somatostatin in EC cells and D cells, respectively. ECL cells, EC cells, P cells, D1 cells and X cells contained CGA in their granules, whereas D cells did not. CGA granular content in ECL cells, P cells, D1 cells and X cells was 3.39 +/- 0.17, 3.41 +/- 0.21, 3.58 +/- 0.18, and 3.55 +/- 0.09, respectively. In ECL cells, CGA was also found in a nongranular form. The CGA content in EC cells (2.95 +/- 0.21) was not significantly different from the serotonin content (2.82 +/- 0.11; p > 0.05) which is in line with the basic significance of CGA as potential amine storage and release protein. The somatostatin content in D cells was 3.30 +/- 0.15. Our study has established high content of CGA in granules of all types of endocrine cells in human oxyntic gastric mucosa except in D cells.

Light and electron microscopic immunohistochemical investigation on G and D cells in antral mucosa in Helicobacter pylori-related gastritis.

Recently, it has been recognized that Helicobacter pylori (H. pylori) infection is associated with an exaggeration of basal and meal gastrin secretion. We investigate whether there is a relationship between H. pylori-related chronic gastritis and G-cell and D-cell number and granule density index of G and D cells. - The number of antral G cells and D cells and granule density index of D and G cells are compared between thirty two patients with H. pylori-related chronic gastritis and twelve patients without H. pylori and inflammation. Antral mucosal biopsy specimens are examined using light and electron immunohistochemical techniques. - The number of G cells is the same in either infected or uninfected patients (98.40 +/- 11.39, 109.25 +/- 12.76 vs 101.17 +/- 7.72 for infected patients with non atrophic and with mild atrophic chronic gastritis and uninfected controls, respectively) except for the cases with moderate gastric mucosal atrophy, where G cells (58.22 +/- 5.63) decrease in number. The number of D cells is decreased in all patients with H. pylori-related gastritis. G cell granule density index is significantly (p < 0.05) increased in patients with H. pylori-related chronic gastritis than in controls (3.15 +/- 0.43 vs 2.528 +/- 0.01). D cell granule density index is similar between patients with H. pylori chronic gastritis and controls (3.18 +/- 0.05 vs 3.166 +/- 0.12). It is concluded that decreased D cells number in patients with H. pylori-related chronic gastritis might be one of the reasons for the existing hypergastrinaemia.

The ultrastructure of primary cilia in the endocrine and excretory duct cells of the pancreas of mice and rats.

A primary cilium was frequently observed in the endocrine alpha, beta and delta cells, as well as in the excretory duct cells of the pancreas of normal mice and rats. The characteristic components of the cilium including the basal body, axoneme (shaft), and terminal part were clearly recognizable. The basal body or distal centriole surrounded by Golgi vesicles was perpendicularly oriented to the proximal centriole, and a dense striated band was seen filling the gap between them. The microtubules of the basal body consisted of nine peripheral triplets exhibiting a 9 + 0 pattern, an appearance similar to that of the proximal centriole. Rootlets, basal feet and alar sheets associated with the basal body were occasionally seen. The axoneme usually consisted of a 9 + 0 pattern of microtubule doublets, but other irregular patterns of 7 + 2, 7 + 3, and 8 + 1 were also seen. The microtubules in the terminal part of the cilium became fewer in number and had no peculiar arrangement. The cilium of the endocrine cells always projected into the intercellular canaliculus and was covered by the ciliary sheath, and occasionally, double cilia were visualized in the vicinity of beta cells. In the excretory duct cells, the cilium showed similar features, but it was slightly longer and always projected into the dense secretory content of duct lumen. On the other hand, no primary cilium was ever observed in the acinar cells of mouse and rat pancreas. In conclusion, the present study describes the morphology of primary cilia and its associated components in the endocrine and excretory duct cells of the pancreas of mice and rats. The findings suggest that the primary cilium should be considered as a constant intracellular organelle though its function and significance remain speculative.

Glucokinase is located in secretory granules of pancreatic D-cells.

We immunohistochemically examined the distribution of glucokinase in rat pancreatic islets. Glucokinase immunoreactivity under light microscopy was detected in the cytoplasm of somatostatin cells as well as in that of insulin cells. No specific immunoreactivity was detected in glucagon and pancreatic polypeptide cells. In somatostatin cells, glucokinase immunoreactivity was located by electron microscopy exclusively within secretory granules.