Normal Rat Pancreas Research Articles

Among Other Tissues, Short-Term Garlic Oral Treatment Incrementally Improves Indicants of Only Pancreatic Islets of Langerhans Histology and Insulin mRNA Transcription and Synthesis in Diabetic Rats.

The source, mRNA transcription, and synthesis of insulin in the pancreas, in addition to the bile duct and liver, in streptozotocin (STZ)-induced diabetic rats (DR) in response to garlic oral treatment are not yet clear. This study investigated the accumulative effects of continued garlic oral treatment on changes in the pancreas, bile duct, and liver with regards to: 1-Insulin mRNA transcription, synthesis, and concentration in relation to changes in serum insulin (SI); 2-Insulinogenic cells insulin intensity and distribution, proliferation, and morphology. Fasting blood glucose (FBG) and insulin concentration in serum and pancreas (PI) and sources and mRNA transcription in the pancreas, bile duct, and liver in normal rats given normal saline (NR-NS) and DR given either NS (DR-NS) or garlic extract (DR-GE) before and after 1, 4, and 8 weeks of oral treatment were examined. Compared to NR-NS, DR-NS showed a significant increase in FBG and reductions in SI and PI and deterioration in islets histology, associated pancreatic insulin numerical intensities, and mRNA transcription. However, compared to DR-NS, the targeted biochemical, histological, and genetic variables of DR-GE were significantly and incrementally improved as garlic treatment continued. Insulin or its indicators were not detected either in the bile duct or the liver in DR-GE. 8 weeks of garlic oral treatment is enough to incrementally restore only pancreatic islets of Langerhans insulin intensity and insulinogenic cells proliferation, morphology, and distribution. These indices were associated with enhanced pancreatic insulin mRNA transcription and synthesis. Eight weeks of garlic treatment were not enough to stimulate insulinogenesis in either the bile duct or the liver.

Serotonin transporter imaging agent as a probe for β-cells of pancreas

ObjectiveDiabetes mellitus (DM) is one of the major diseases in the world. Nuclear medicine imaging may be able to detect functional status of pancreatic β cells in vivo, which might elucidate the pathological mechanisms of diabetes and develop individualized treatment plans. In this study, we evaluated the ability of [125I]ADAM, a serotonin transporter (SERT) imaging agent, as a probe for detecting pancreatic β-cell mass (BCM). MethodsIn vitro cell studies were evaluated in INS-1 cells (rat islet β cell line). Biodistribution studies were performed in male normal Sprague-Dawley rats and alloxan-induced type 1 diabetes mellitus (T1DM) rats. Distribution and expression of SERT protein in pancreas of rats were also measured by immunofluorescence staining and Western blot. ResultsIn vitro cell studies showed that the concentration of [125I]ADAM associated with the INS-1 cells was increased gradually with incubation time, and the SERT specific inhibitor, escitalopram, exhibited the inhibitory effect on this interaction. Biodistribution studies also showed that the uptake of [125I]ADAM in the pancreas of normal rats was decreased in the presence of escitalopram. However, in the T1DM rat model with a significant β cells reduction, the uptake of pancreas was increased when compared with the control. Through immunofluorescence staining and Western blot, it was found that both the endocrine and exocrine cells of the normal pancreas expressed SERT protein, and the level of SERT protein in the exocrine cells was higher than islets. In the diabetic state, the expression of SERT in the exocrine cells was further increased. ConclusionsThe SERT imaging agent, [125I]ADAM, at the present form will not be suitable for imaging β cells, specifically because there were extraordinarily high non-specific signals contributing from the exocrine cells of pancreas. In addition, we noticed that the level of SERT expression was abnormally elevated in the diabetic state, which might provide an unexpected target for studying the pathological mechanisms of diabetes.

Spexin Co‐localizes with Insulin in Pancreatic Islet Cells of Normal and Diabetic Rats

Spexin is a newly discovered gastrointestinal tract (GIT) peptide. Preliminary biochemical analysis showed that spexin was localized to secretory granules in a transfected pancreatic cell line. Spexin was also expressed in the submucosal cells of the mouse stomach indicating that it is a resident peptide in the GIT and may thus influence pancreatic function. It has been suggested that spexin may play a role in the regulation of physiological homeostasis. Our aim was to determine whether spexin is present in the pancreas of normal and streptozotocin‐induced diabetic rats. We also wanted to know whether spexin co‐localizes with pancreatic hormones in the islet of Langerhans. Immunohistochemical, immunofluorescence and transmission electron microscopy (TEM) methods were used to determine the cellular localization of spexin in the endocrine pancreas of non‐diabetic and diabetic rats. Immunofluorescence study showed that a significant number of cells in the core of rat pancreatic islet contains spexin, where insulin‐positive cells are located. We used double labelling immunofluorescence method to determine whether spexin and insulin co‐localize together in the same cell. We observed that these spexin‐immunoreactive cells also contain insulin. There was no evidence of co‐localization of spexin with either somatostatin or pancreatic polypeptide. The number of spexin‐ and insulin‐immunopositive cells was significantly (p <0.05) reduced after the onset of diabetes mellitus. Moreover, TEM showed that the secretory granules of pancreatic beta cells contain spexin‐labelled immunogold particles. In conclusion, the presence of spexin in pancreatic beta cells suggests a role in the regulation of pancreatic endocrine function.

Palm leaf catechins improved insulin-related pathways in diabetic rats

Catechin-rich oil-palm leaf extract (OPLE) (Elaeis guineensis) was previously demonstrated to possess benefits for diabetes and cardio metabolic health (vasodilation, antioxidant, cardiovascular, anti-hypertensive, anti-inflammatory, hepatoprotective and nephroprotective properties) in animal models. For insights into OPLE anti-diabetic mode-of-action and possible toxicity, the effects of dietary OPLE on insulin-signaling pathways mRNA expressions in the liver, kidney, pancreas, and spleen of normal and diabetic rats were examined. Type-2-Diabetes Mellitus (T2DM) were induced by chronic high-fat diet and streptozotocin (35 mg/kg) intraperitoneal injection. The OPLE (100 mg/kg body weight) were fed daily to normal and T2DM-induced rats. The OPLE suppressed hyperglycaemia and excessive weight gain in the T2DM rats, and appeared harmless to normal rats. The OPLE supplementation significantly (p<0.05) modulated the mRNA expressions of phosphatidylinositol-3 kinase (PIK3R1); insulin signaling receptor (INSR); insulin-receptor substrates 1 and 2; and ectonucleotide pyrophosphatase-1 (ENPP1) especially in the livers of normal rats and the spleen of diabetic rats. Results suggested the OPLE probably help prevent diabetes in healthy mammals and ameliorate the immune functions of diabetic mammals. The OPLE improved the antioxidant defence responses, insulin-pathways mRNA expressions in the normal and diabetic rats; suppressed hyperglycaemia and excessive weight gain in T2DM rodents without observable liver or kidney toxicity at the dose used.

Antidiabetic and antihyperlipidemic activities of a novel polyherbal formulation in streptozotocin - nicotinamide induced diabetic wistar rats

Objective: To investigate the antidiabetic and antihyperlipidemic activities of polyherbal formulation (PHF) containing seven plants namely Cassia auriculata, Cassia fistula, Syzygium cumini, Cyperus rotundus, Saussurea lappa, Terminalia arjuna and Salacia reticulate in streptozotocin (STZ) - nicotinamide (NC) induced diabetic rats by administering oral doses (200 and 400 mg/kg body weight).
 Materials and Methods: Animals were divided into diabetic and nondiabetic groups. Rats were fed with normal laboratory diet and induced with a single intraperitoneal injection of 60mg/kg of STZ, and thereafter 120 mg/kg NC was injected after 15min. Diabetic rats were treated with formulation (200 and 400 mg/kg) and glibenclamide 5 mg/kg. Blood glucose levels were measured using blood glucose test strips with ACCU CHEK glucometer. Glycosylated haemoglobin, total haemoglobin, lipid profiles, lipoproteins, hepatic marker enzymes activity were determined in normal and STZ- NC induced diabetic rats after oral administration of the PHF for 28 days. Histopathological changes in normal and diabetic rat pancreas organs were also observed after PHF treatment.The statistical analysis of results was carried out using one-way analysis (ANOVA) followed by post hoc multiple comparison tests.
 Results: Treatment of diabetic rats with PHF (200 and 400 mg/kg) and glibenclamide (5 mg/kg) indicate significant decreased blood glucose level and significant improvement in body weight. PHF treated rats showed significant (P < 0.01) decrease in the level of HbA1C, TC, TG, LDL, VLDL, AST, ALT and ALP while a significant increment in the level of Hb, HDL cholesterol was observed. Furthermore, the PHF treated rats has a favourable effect on the histopathological changes of the pancreas in STZ-NC induced diabetes.
 Conclusion: These findings suggested the antihyperglycemic and antihyperlipidemic properties of the PHF and thus help in preventing future complications of diabetes.

Phytochemical and Hypoglycemia Effect of Tomato Lycopene Extract (TLE)

Background: High blood glucose level is one of the Diabetes Mellitus (DM) symptoms. DM type 2 occurs when the body's cells experience insulin resistance. Insulin resistance caused by an increase of lipids in the body, so insulin can not be used properly. Objective: To determine the effect of giving tomato lycopene extract (TLE) to reduce levels of rat blood glucose induced by hyperglycemia. Method: Tomato skin extracted with four kinds of solvents, namely acetone, ethanol, n-hexane, and a combination of n-hexane: acetone: ethanol 96% (1: 2: 1). The identification of lycopene compounds was carried out by the TLC derivatization method. Hyperglycemia rats were induced by a high-fat diet (HFD) for 30 days and dexamethasone 1 mg/kg BW (i.p). Blood glucose levels were measured before and after the administration of the extract. TLE 5, 15, and 50 mg/kg BW given for 14 days. Results: The combination of n-hexane: acetone: ethanol 96% (1: 2: 1) produces the most lycopene content. This extract contains the fewest phenol and flavonoid compounds, so it is called a tomato lycopene extract (TLE). LTE 5, 15, and 50 mg/kg BW can reduce blood glucose levels. On the 7th and 14th day, the rat blood glucose level of the treatment group was not statistically different from the normal group. Conclusion: The hyperglycemic rat pancreas did not show any significant difference to the pancreas of normal rats. TLE 5, 15, and 50 mg/kg BW can reduce blood glucose levels.

2156-P: Development of the Immune System in the Normal Rat Pancreas

The role of resident immune cells in organogenesis has not been extensively studied in pancreas. We hypothesized that during the critical stage of ß-cell remodeling during the newborn period, immune cell populations would also change. Using flow cytometry and immunohistochemistry, we identified immune cell populations in the endocrine (islets) and exocrine portion of the pancreas at postnatal day 1, 7 and 14. When corrected for the expected increase in pancreatic weight, immune cell populations increase in the exocrine pancreas in males and females from P1 to P7 but not further at P14. Interestingly, in islets the number of immune cells further increases in males from P7 to P14. Macrophages, T cells, B cells, granulocytes and NK cells are present in islets and exocrine pancreas. Macrophages are the dominant population in islets (M: 47%, F: 47%) and exocrine (M:30%, F: 34%) pancreas. In islets, M2 macrophages are favored at P1. M1 macrophages dramatically decrease with age. In exocrine pancreas, M1 macrophages are initially favored, but with age the M2 population increases. T cells are also present in islets (M:6%, F:6%) and exocrine (M:8%, F: 6%) pancreas. Cd3+ Cd4- Cd8- is the dominant T cell population at P1 but decreases with age replaced by Cd3+ Cd4+ Cd8- cells. In islets from male, but not females, the number of T cells, specifically Cd4+ T cells, increases with age. B cells (Cd45Ra+) are present in islets (M:20%, F:19%) and exocrine (M:20%, F:25%) pancreas and numbers do not change with age. Neutrophils (HIS48hi Cd68) are also present in islets (M:6%, F:6%) and exocrine (M:13%, F: 10%) pancreas. NK cells are a minor cell population present at about 1% of total immune cells in male and female pancreas at P1. Interestingly, NK cells increase with age in females in islets and exocrine pancreas. These data demonstrate significant sex specific changes in the pancreas during the critical period of postnatal pancreas development suggesting a role for immune cells in normal postnatal development and function in the endocrine and exocrine pancreas. Disclosure T. Golden: None. G. Worthen: None. R.A. Simmons: None. Funding National Institutes of Health

Primary Cultures for Pancreatic Stellate Cells (PSCs).

Primary cultures of pancreatic stellate cells (PSCs) remain an important basis for in vitro study. However, effective methods for isolating abundant PSCs are currently lacking. This purpose of this chapter is to report our novel approach to isolating PSCs from normal rat pancreas and human pancreatic ductal adenocarcinoma (PDAC) tissue. Normal PSCs were isolated with enzyme digestion and ladder centrifugation with Nycodenz solution. Isolated PSCs were cultured in DMEM/F12 containing 10% fetal bovine serum. Cancer-associated PSCs were obtained by an outgrow method from fresh human PDAC tissues. Isolated activated PSCs were cultured in DMEM/F12 containing 20% fetal bovine serum. With our modification, normal pancreas tissue yields an adequate number of PSCs (approximately 0.5-5million/g pancreas) for in vitro studies, and the cell viability was about 90%. And a modified outgrowth method made tissue blocks attached more tightly and significantly shortened the outgrowth time of the activated cells. Our modification in PSC isolation methods significantly increased the isolation efficiency and shortened the culture period, thus facilitating future PSC-related research.

Glucose Lowering Activity of the Aqueous Extract of Warionia saharae in Normal and Diabetic Rats.

Warionia saharae is a medicinal plant used in the Morocco for the treatment of diabetes mellitus. The aim of the study was to assess the effect of the aqueous extract of this plant on blood glucose levels in diabetic rats. The current study was carried out to investigate the effect of a single dose and daily oral administration for 15 days of the Aerial Part Aqueous Extract (A.P.A.E) of Warionia saharae (W. saharae) at a dose of 5 mg/kg body weight on blood glucose levels and the histopathological changes in liver and pancreas both in normal and streptozotocin-induced diabetic rats (STZ). Also, the effect of this extract to improve glucose tolerance in normal rats was demonstrated. Single oral administration of W. saharae A.P.A.E reduced blood glucose levels 6 h after administration in normal rats and in STZ-induced diabetic rats. Furthermore, blood glucose levels were decreased in normal and STZ-induced diabetic rats after 15 days of treatment. The aqueous extract of W. saharae was shown to improve the increase on blood glucose levels in normal treated rats significantly (p< 0.001) 90 min after glucose administration as compared to the control groups. In conclusion, the results show that APAE of W. saharae possesses significant antihyperglycemic and hypoglycemic activity.

Effect of nociceptin on insulin release in normal and diabetic rat pancreas.

Nociceptin (NC), also known as Orphanin FQ, is a brain peptide involved in the regulation of pain, but its role in the endocrine pancreas is poorly understood. The present study examines the pattern of distribution of NC and its effect on insulin and glucagon secretion after the onset of diabetes mellitus (DM). Male Wistar rats weighing 150-200g were made diabetic with streptozotocin (60mg/kg body weight, intraperitoneally). Four weeks after the induction of DM, pancreatic tissues were retrieved and processed for immunofluorescence, immunoelectron microscopy, and insulin and glucagon secretion. Isolated islets from non-diabetic and diabetic rats were used to determine the effect of NC on insulin release. NC was discerned in islet cells of non-diabetic control and diabetic rat pancreata. NC co-localized only with insulin in pancreatic beta cells. NC did not co-localize with either glucagon or somatostatin or pancreatic polypeptide. The number of NC-positive cells was markedly (p < 0.001) reduced after the onset of DM. Electron microscopy study showed that NC is located with insulin in the same secretory granules of the beta cells of both non-diabetic and diabetic rat pancreas. NC inhibits insulin release markedly (p < 0.05) from pancreatic tissue fragments of non-diabetic and diabetic rats. In contrast, NC at 10-12M stimulates insulin release in isolated islets of DM rats. In conclusion, NC co-localizes with insulin only in the islet of Langerhans. The co-localization of NC with insulin suggests a role for NC in the regulation of pancreatic beta cell function.

Hepatic, Pancreatic, and Renal Histo-Morphologic Alterations in Administration of Aqueous and Ethanol Seed Extract of Buchholzia Coriacea in Alloxan-Induced Diabetic Rats

Physiologically, Insulin is a hormone that helps the body’s cells to absorb glucose (sugar) so it can be used as a source of energy. In diabetics however, due to pancreatic production of insufficient amounts of insulin or failure of body’s cells to respond to available insulin, blood and urine glucose levels build up to cause excessive urination, thirst, hunger, and problems with fat and protein break down. Recently, medicinal herbs have been implicated in traditional medical practice for the treatment of this ailment (Diabetes Mellitus, DM). Present study sought to investigate the effect(s) of oral administration of aqueous and ethanol extracts of Buchholzia Coriacea on the liver, pancreas and kidney in normal and alloxan-induced diabetic rats. Forty adult rats of both sexes were randomly assigned into two groups (normoglycemic and hyperglycemic). While group 1 (normoglycemic) had normal control, metformin, aqueous extract (250 mg/kg) and ethanol extract (250 mg/kg) treated sub-groups respectively, Group 2 (hyperglycemic) contained the diabetic control, metformin, aqueous extract (250 mg/kg), and ethanol extracted (250 mg/kg) treated sub-groups dosed daily by oral gavage for 14 days. At the end of treatment, rats were euthanized via cervical dislocation; with selected visceral (liver, pancreas and kidney) harvested and observed for histo-architectural changes. Blood samples were also collected and checked for sugar levels. Slight modulations were seen in the histo- morphology of the pancreas, liver and renal tissues as Buccholzia Coriacea apparently posed some hypoglycemic effects. Hitherto, there was an appreciable improvement and merit in the use of the extract in the management of diabetes across groups.

Antidiabetic effect of Ruta montana L. in streptozotocin-induced diabetic rats.

Ruta montana (L.) is known as a medicinal plant with many beneficial effects, including those that can be used in the treatment of diabetes. The objective of the study was to investigate the antidiabetic effect of this plant in diabetic rat. This study investigated the effects of an aerial part aqueous extract (APAE) of Ruta montana (L.) (RM) at a dose of 5 mg/kg on blood glucose levels in normal and streptozotocin (STZ)-induced diabetic rats. Histopathological changes were also evaluated in liver and pancreas both in normal and STZ-induced rats. The effect of this aqueous extract on glucose tolerance was demonstrated in normal rats. Furthermore, the relative organ weight (ROW) of liver, kidney, pancreas, and brown adipose tissue were evaluated after 15 days of daily oral administration of the aqueous extract. Both single and repeated oral doses of APAE (5 mg/kg) produced significant reductions in the blood glucose levels in normal and STZ-induced rats. Oral glucose tolerance test results showed that, after the administration of 3 g/kg of glucose, RM APAE (5 mg/kg) improved the increase in blood glucose levels in normal rats at the 30th min (p<0.01) and 90th min (p<0.001). RM APAE exhibits a potent hypoglycemic effect in normal rats and an antidiabetic effect in STZ-induced rats. This finding supports the use of this plant in traditional Moroccan medicine for diabetes management.

Antioxidant and Postprandial Glucose-lowering Potential of the Hydroethanolic Extract of Nypa fruticans Seed Mesocarp

Postprandial hyperglycemia and oxidative stress are implicated in the pathophysiology of type 2 diabetes. An alternative treatment should target both conditions. The present study aimed to evaluate the antioxidant and postprandial glucose-lowering potential of a polyphenolic rich extract of Nypa fruticans nuts. Hydroethanolic extract (HEE) was prepared; polyphenol and fl avonoid content was determined. In vitro antioxidant activity was done through molybdenum (VI) reduction and ABTS and DPPH radical-scavenging capacities. The glucose-lowering potential of the extract was evaluated through in vitro glucose-binding capacity, the regulatory ability of the extract on the pancreas of normal Wistar rats (oral glucose tolerance test), and digestion of carbohydrates (starch and sucrose) by normal Wistar rats. Starch (1 g/kg of the body weight (BW) and sucrose (2 g/kg BW) solution were used as substrates for alpha amylase and invertase inhibition test, respectively. Acarbose (3 mg/kg BW) was used as the reference. Polyphenolic-rich extract (400 mg/kg BW) was administered to treated rats. Results showed that HEE of N. fruticans nuts has antioxidant activity through the scavenging of ABTS (EC50 of 1.75 mg/mL) and DPPH (2.96 mg/mL) radicals and the reduction of Mo (VI). The extract signifi cantly reduced glucose concentrations at t30 compared to glibenclamide, but glibenclamide maintained lower glycemia than the extract thereafter to the end of the study. From the administration of sucrose, the extract maintained lower plasma glucose levels compared to acarbose throughout the study. However, no inhibition was observed in glycemia following starch administration in HEE-treated group as glycemia remained higher compared to the acarbose group throughout the study. Thus, N. fruticans seed mesocarp is a rich source of dietary polyphenols with a strong antioxidant capacity. Its HEE has glucose-lowering potential and possible inhibitory action on the last phase of carbohydrate digestion.

Investigation of an antidiabetic potential of a phenolic compound isolated from the leaves of mempelas paya

Introduction: Leaves of mempelas paya (Tetracera indica) (family: Dilleniaceae) is traditionally used in diabetes management in Malaysia. However, no attempt has ever been made to discover an active principle responsible for its blood glucose lowering effect. Hence, this research deals with the isolation, in vivo antidiabetic and toxicological evaluation of an active principle. Methods: Chemical investigation of methanol extract derived from the powdered leaves resulted in the isolation of a phenolic compound through repeated silica gel column chromatography. The structure was elucidated by spectroscopic (1H-NMR, 13C-NMR, MS, IR and UV) analysis. In vivo antidiabetic and toxicological studies were carried out using Sprague-Dawley rats (diabetic and normal) at three different concentrations (1, 5 and 25 mg/kg b.w.). Results: 5,7-dihydroxy-8methoxyflavone was found to exhibit significant anti-hyperglycaemic activity (p&lt;0.05) in diabetic rats at 5 and 25 mg/kg b.w. without showing hypoglycaemic effect in normal rats. The anti-hyperglycaemic activity was found to be comparable with glibenclamide (GLBC) (p&lt;0.05). In vivo toxicological study for continuous 21 days showed no abnormal activity and mortality. Histopathology of kidney, liver and pancreas of both normal and diabetic treated rats demonstrated normal and improved condition when compared to diabetic control group. There were no significant changes seen in the kidney (serum urea and creatinine) and lipid (TC, TG, LDL-C and HDL-C) functional parameters tested. Conclusions: 5,7-dihydroxy-8-methoxyflavone isolated from the leaves of mempelas paya coud be a safe antidiabetic agent.

Diabetes Mellitus Influences the Pattern of Distribution of Irisin in Pancreatic Islet Cells

Irisin is a newly discovered myokine, which enhances the turnover of white adipose cells into brown fat cells. Irisin is derived from FNDC5, a membrane‐bound protein. Moreover, irisin has been shown to play an important role in energy regulation especially in muscle cells. Although, irisin has been detected in many organs including the exocrine pancreas, liver, testes and sebaceous gland, a precise localization of this polypeptide in pancreatic islet cells has not been undertaken. Immunohistochemistry was used to determine the pattern of distribution of irisin in streptozotocin‐induced (60 mg/Kg body weight) diabetes in rats. Image J® software was used to quantify the number of irisin‐immunoreactive cells in diabetic and non‐diabetic rat pancreas. Our study shows that irisin is present in a large number of pancreatic beta cells where it co‐localizes with insulin. In contrast, irisin does not co‐localize with glucagon in normal rat pancreas. However, this pattern of distribution changes after the onset of diabetes mellitus (DM). DM is characterized by increased number of glucagon immunopositive cells. The newly formed glucagon‐positive cells now contain irisin as well. The reason why irisin co‐localizes with glucagon in diabetic rat pancreas and not in non‐diabetic rats is unclear. It is possible that in diabetic rats, insulin, the natural inhibitor of glucagon has been removed paving way for glucagon expression in pancreatic beta cells devoid of insulin. The presence of irisin in insulin secreting beta cells suggests that the beneficial effect of irisin on glucose metabolism could indeed be via the endocrine pancreas.Support or Funding InformationUnited Arab Emirates University

C4-P-10Three-dimensional ultrastructural analysis of secretory granules in histological paraffin sections of endocrine tumors using low-vacuum SEM

Secretory granules of endocrine tumors are traditionally identified by transmission electron microscopy of tissue samples. Recently, we developed a simple method for three-dimensional morphological analysis of conventional paraffin sections using low-vacuum scanning electron microscopy (SEM) [1], and showed that this technique was useful for pathological diagnosis of renal biopsy samples [2]. In this study, we investigated whether this method is useful as an adjunct in pathological diagnosis of endocrine tumors. Preparations were processed according to our original method. Paraffin sections (5 μm thick) obtained from a normal rat pancreas as a control, a human pituitary adenoma and a colorectal carcinoid tumor were generated by the conventional method. After deparaffinization, the sections were stained only with platinum blue aqueous solution (TI-blue staining kit: Nisshin EM, Tokyo) for 15 min at room temperature, and then observed by low-vacuum SEM (TM-1000, TM3030 plus: Hitachi, Tokyo) at high magnification (~30,000×). Under low-vacuum SEM, secretory granules of each cell could be identified as bright spherical granules in each paraffin-sectioned tissue, depending on the contrast of BSE signals. Figure shows low-vacuum SEM image (a) and TEM image (b) of endocrine secretory granules 100–260nm in diameter (arrows) in colorectal carcinoid tumor cells. Our results suggest that low-vacuum SEM of conventional paraffin sections is suitable for the pathological diagnosis of endocrine tumors. [1] Inaga et al., Arch. Histol. Cytol., 72 (2009) 101-106. [2] Okada et al., Biomed. Res., 35 (2014) 345-350. C4-P-09 doi:10.1093/jmicro/dfv327

Investigation of In vitro PDT Activities and In vivo Biopotential of Zinc Phthalocyanines Using 131I Radioisotope

Novel octylthio-containing asymmetrically substituted Zn(II) phthalocyanine (Zn(II)Pc1) and a symmetric derivative (Zn(II)Pc2) have been prepared to investigate the biological potential and ability to photosensitize singlet oxygen for photodynamic therapy applications. In this study, the singlet oxygen generation potential and in vitro photodynamic activities of these compounds have been tested. Both ZnPcs reveal to be very efficient singlet oxygen generators and promising PSs for PDT applications. In vitro PDT activities of the compounds were evaluated in EMT-6 murine mammary carcinoma and HeLa human cervix carcinoma cell lines. Moreover, Zn(II)Pc1 displayed the phototoxic effects in the mammary cancer cell line (6.25 μm concentration at 30 J/cm(2) light dose and 12.5 μm concentration at 20 J/cm(2) light dose), while Zn(II)Pc2 did not show any phototoxic effects both in two cell lines. Zn(II)Pcs were radiolabeled with (131) I in high yields. Biodistribution studies revealed that the radiolabeled Zn(II)Pc1 showed significant uptake in l. intestine, pancreas, brain, and ovary, while Zn(II)Pc2 has significant uptake in ovary and pancreas in normal rats. Hence, these Pcs derivatives could be promising candidate for tumor nuclear imaging.

Generation of functional organs from pluripotent stem cells

Hematopoietic stem cells (HSCs) have played a major role in stem cell biology, providing many conceptual ideas and models. Among them is the concept of the "niche", a special bone-marrow microenvironment that by exchanging cues regulates stem-cell fate. The HSC niche also plays an important role in HSC transplantation. Successful engraftment of donor HSCs depends on myeloablative pretreatment to empty the niche. The concept of the stem-cell niche has now been extended to the generation of organs. We postulated that an empty "organ niche" exists in a developing animal when development of an organ is genetically disabled. This organ niche should be developmentally compensated by blastocyst complementation using wild-type primary stem cells (PSCs). We proved the principle of organogenesis from xenogeneic PSCs in an embryo unable to form a specific organ, demonstrating the generation of functionally normal rat pancreas by injecting rat PSCs into pancreatogenesis-disabled mouse embryos. This principle has held in pigs. When pancreatogenesis-disabled pig embryos underwent complementation with blastomeres from wild-type pig embryos to produce chimeric pigs, the chimeras had normal pancreata and survived to adulthood. Demonstration of the generation of a functional organ from PSCs in pigs is a very important step toward generation of human cells, tissues, and organs from individual patients' own PSCs in large animals.

Histopathological changes in rat pancreas and skeletal muscle associated with high fat diet induced insulin resistance

The effects of a high fat diet on the development of diabetes mellitus, insulin resistance and secretion have been widely investigated. We investigated the effects of a high fat diet on the pancreas and skeletal muscle of normal rats to explore diet-induced insulin resistance mechanisms. Forty-four male Wistar rats were divided into six groups: a control group fed standard chow, a group fed a 45% fat diet and a group fed a 60% fat diet for 3 weeks to measure acute effects; an additional three groups were fed the same diet regimens for 8 weeks to measure chronic effects. The morphological effects of the two high fat diets were examined by light microscopy. Insulin in pancreatic islets was detected using immunohistochemistry. The homeostasis model assessment of insulin resistance index and insulin staining intensity in islets increased significantly with acute administration of high fat diets, whereas staining intensity decreased with chronic administration of the 45% fat diet. Islet areas increased significantly with chronic administration. High fat diet administration led to islet degeneration, interlobular adipocyte accumulation and vacuolization in the pancreatic tissue, as well as degeneration and lipid droplet accumulation in the skeletal muscle tissue. Vacuolization in the pancreas and lipid droplets in skeletal muscle tissue increased significantly with chronic high fat diet administration. We suggest that the glucolipotoxic effects of high fat diet administration depend on the ratio of saturated to unsaturated fatty acid content in the diet and to the total fat content of the diet.

Distribution of nociceptin in pancreatic islet cells of normal and diabetic rats.

Nociceptin has been reported to play an important role in the regulation of pancreatic exocrine secretion. Most of the studies performed on nociceptin are mainly physiological rather than morphological in nature. The present study investigated the pattern of distribution of nociceptin in the endocrine pancreas of normal and diabetic rats. Immunohistochemistry, immunofluorescence, Western blot, and double-labeled immunoelectron microscopy were used in this study. Diabetes was induced using streptozotocin (60 mg/kg body weight). Nociceptin-immunoreactive cells were observed in the central and peripheral regions of the islets of both normal and diabetic rat pancreas. The number of nociceptin-positive cells was significantly (P < 0.05) lower in the islet of diabetic rats compared with the control. Immunofluorescence study showed that nociceptin colocalizes with insulin in pancreatic β-cells. The degree of colocalization of nociceptin with insulin was severely deranged after the onset of diabetes. Moreover, immunogold particles conjugated with either nociceptin or insulin were observed on the granules of pancreatic β-cell. The number of nociceptin-labeled colloidal gold particles was significantly lower after the onset of diabetes. Nociceptin is present in pancreatic islets cells and colocalizes with insulin. Nociceptin may have a physiological role in the metabolism of insulin.