Studies on colocalization of neuropeptide Y,vasoactive intestinal polypeptide,catecholamine-synthesizing enzymes and acetylcholinesterase in the larynx of the rat

Summary In the present immunohistochemical study, the distribution of nerve fibers containing neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP) in the larynx was examined and compared with that of fibers containing tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (BDH), and with that of acetylcholinesterase (AChE)-positive nerve fibers, in intact and vagotomized rats and in rats subjected to removal of the superior cervical ganglion (SCG). Fibers showing TH/DBH-like immunoreactivity (LI) were only found in the walls of arteries and arterioles, whereas AChE-positive nerve fibers were located close to the acini and ducts of the glands, in blood vessel walls, in the perichondrium and in the lamina propria. NPY-LI and VIP-LI coexisted in local AChE-positive ganglionic cells and in a subpopulation of the AChE-positive fibers, NPY-LI also being present in some periarterial fibers showing TH/DBH-LI. Unilateral removal of the SCG eliminated the TH/DBH-innervation in the upper but not the lower parts of the larynx ipsilaterally, whereas the NPY-innervation of the arteries in the upper parts only partly disappeared and the NPY-innervation of the other structures remained unchanged. The distribution of VIP-innervation was unchanged after vagotomy and removal of the SCG. The results suggest that VIP is present in the postganglionic parasympathetic innervation, whereas NPY is present in both the postganglionic parasympathetic and sympathetic innervation of the rat larynx.     

Distribution and origin of vasoactive intestinal polypeptide (VIP)-immunoreactive,acetylcholinesterase-positive and adrenergic nerves of the cerebral arteries in the bent-winged bat (Mammalia: Chiroptera)

Summary The overall distribution and origins of vasoactive intestinal polypeptide (VIP)-immunoreactive (IR), acetylcholinesterase (AChE)-positive and adrenergic nerves in the walls of the cerebral arteries were investigated in the bent-winged bat. VIP-IR and AChE-positive nerves innervating the bat cerebral vasculature appear to arise mainly from VIP-IR and AChE-positive cell bodies within microganglia found in the nerve bundle accompanying the sympathetic nerve bundle within the tympanic cavity. These microganglia, as well as the nerve bundle containing them, do not emit catecholamine fluorescence, suggesting that they are of the cranial parasympathetic outflow, probably the facial or glossopharyngeal one. The axons from VIP-IR and AChE-positive microganglia run intermingled with sympathetic adrenergic nerves in the same thick fiber bundles, and reach the cranial cavity through the carotid canal. In addition, some of the VIP-IR fibers innervating the vertebro-basilar system, at least the basilar artery, originate from VIP-IR nerve cells located in the wall of this artery.The supply of VIP-IR fibers to the bat major cerebral arteries is the richest among mammals that have been studied, and differs from other mammals in that it is much greater in the vertebro-basilar system than in the internal carotid system: plexuses of VIP-IR nerves are particularly dense along the walls from the posterior ramus to posterior cerebral and basilar arteries. Small pial and intracerebral arteries of the vertebro-basilar system, especially those of the posterior cerebral artery which supply most parts of the diencephalon and cerebrum, are also richly innervated by peripheral VIP-IR fibers. This pattern corresponds well with the innervation pattern of adrenergic and AChE-positive nerves.     

On the intrinsic innervation of normal rat liver

Summary With the Bodian method stained fibers were observed in the lobules of the rat liver and with the modified Karnovsky and Roots thiocholine method cholinesterase (presumably acetylcholinesterase (AChE))-positive nerve fibers were found in a pattern similar to that of the Bodian-stained fibers. The AChE positive nerve fibers form a network in the liver lobules in close relation to hepatocytes and sinusoids. Fluorescent varicose nerve fibers demonstrated by the glyoxylic acid and Falck-Hillarp fluorescence methods were found only in the interlobular spaces associated with vessels. As no overlapping of distribution patterns of AChE-positive nerve fibers and fluorescent nerve fibers occurs, the AChE activity of the nerves of the liver lobules probably reflects the associated presence of acetylcholine in the nerve fibers. In consequence we suggest that nerves of the liver lobules belong to the autonomic parasympathetic nervous system.SEM of liver tissue revealed light cords apparently situated in smooth surfaced channels between adjacent hepatocytes and in the space of Disse, where fibers also cross sinusoids. We tentatively suggest that the cords of the SEM represent the AChE-positive nerve fibers of our LM observations'.The skilled assistance of Dr. Esther Hage, Department of Pathology, Odense Sygehus, Denmark, in the Falck-Hillarp fluorescence work is gratefully acknowledged     

Enkephalin-like immunoreactivity in ganglionic cells in the larynx and superior cervical ganglion of the rat.

The distribution of enkephalin-like immunoreactivity (ENK-LI) in the larynx, the superior cervical ganglion (SCG) and the nodose ganglion of adult rats was examined in the present study. A substantial number of the local acetylcholinesterase (AChE)-positive, presumably parasympathetic, ganglionic cells in the larynx displayed ENK-LI. These cells also exhibited neuropeptide Y (NPY)- and vasoactive intestinal polypeptide (VIP)-LI. Varicose nerve fibers showing ENK-LI were observed close to the acini and ducts of the glands, in the perichondrium and in the lamina propria. The varicosities exhibiting ENK-LI frequently displayed NPY- and VIP-LI. The ENK-LI was detected in a subpopulation of AChE-positive nerve fibers in the laryngeal tissue. In the SCG, only a small number of the ganglionic cells displayed ENK-LI. These cells, in contrast to other ganglionic cells of the SCG, did not show NPY-LI. None of the ganglionic cells of the nodose ganglion exhibited ENK-LI. Sympathectomy and vagotomy affected neither the number nor the distribution of fibers showing ENK-LI in the larynx. In conclusion, ENK appears to be present together with NPY and VIP in the parasympathetic innervation of the larynx and in a very limited number of the ganglionic cells of a sympathetic ganglion, the SCG, of the adult rat.     

Ultrastructural localization of acetylcholinesterase (AChE) activity in the chicken Harderian gland

Localization of acetylcholinesterase (AChE) was investigated in the chicken Harderian gland at the electron microscopic level. Nerve cells in the pterygopalatine ganglion showed AChE activity. They had a pale and large nucleus which was round or oval in shape. Reaction product of AChE was detected between the nuclear envelopes; in the cisterna of rough endoplasmic reticulum and the lumen of the Golgi lamellae, and on the plasma membrane of the nerve cell. In the interstitium of the gland, nerve fibers showing AChE activity were easily found. They were often seen in the perivascular space and between plasma cells. These nerve fibers had varicosities in contact with plasma cells and the endothelium or the smooth muscle fiber of the blood vessels. AChE-positive varicosities or terminals contained many small clear vesicles (about 50nm in diameter) and a few large dense-cored vesicles (about 100 nm in diameter). No contacts of nerve fibers with acinar cells or the ductal epithelium were observed in the present study. Our data indicate that cholinergic nerves play distinct roles in the regulation of the immune function of the chicken Harderian gland.     

Effect of pesticide bendiocarbamate on distribution of acetylcholine- and butyrylcholine-positive nerves in rabbit's thymus

Many pesticides used in agriculture have a negative effect on organisms. The group of hazardous pesticides includes the cholinesterase inhibitor bendiocarbamate. According to literature, bendiocarbamate has relatively low toxicity in mammals and vertebrates in general, since it does accumulate in their tissues and the cholinesterase activity returns to norm within 24 h after acute exposure. The present study focused on the influence of bendiocarbamate on rabbit thymus after its administration at a dose of 5 mg/kg body weight (BW) for the period of 3 months. The thymus was observed for acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE)-positive nerve fibers visualized by histochemical methods. Microscopic findings of BuChE-positive nerve fibers show the same density and the topography by the experimental and the control animals. On the other hand, AChE-positive nerve fibers in experimental animals after administration of bendiocarbamate is only poorly identified, suggesting that bendiocarbamate inhibits AChE but not BuChE.     

The effect of nerve growth factor on cholinergic neurons in dissociated cultures of the septum pellucidum

Neurons dissociated from septal area of fetal (E 18-19) rat brain were grown for 7 days in culture. Cholinergic neurons were identified by cytochemical demonstration of acetyl cholinesterase. Addition of the nerve growth factor to the culture medium (200 u/ml) increases the number of AChE-positive neurons, the size of the cell body and activity of AChE in these neurons.     

Short-time predegenerated peripheral nerve grafts promote regrowth of injured hippocampal neurites

Our previous studies revealed that predegenerated peripheral nerve grafts facilitated neurite outgrowth from the injured hippocampus and that this effect was particularly distinct when 7-, 28-, and 35-days predegenerated nerve grafts were used. It is recently known that a totally transected peripheral nerve exhibits biphasic neurite-promoting activity. The early phase lasts 7 days. The aim of the present study was to find whether short-time predegenerated (1-6 days) peripheral nerve grafts exert any neurotrophic effect and when this influence is maximal. Experiments were carried out on adult male Wistar rats. Sciatic nerves were totally transected and following 1, 2, 3, 4, 5 and 6 days their distal stumps were implanted into the hippocampus. Control animals were treated with non-predegenerated sciatic nerve grafts. In all groups FITC-HRP was injected into the free end of graft six weeks following surgery. Special histochemic technique showed AChE-positive fibres inside the grafts of all examined groups. Fluorescence microscopic examination revealed the labeled cells in all examined groups, however their number was different in each group, depending on the predegeneration stage. They were most numerous at the fourth day of predegeneration.     

Study of effect of embryonic anlage allografts of the rat spinal cord on growth of regenerating fibers of the recipient nerve

A comparative study of the effect of tissue and suspension allografts of an embryonic spinal cord on regeneration of nerve fibers of impaired (by application of a ligature) sciatic nerve in rats was conducted. It was demonstrated that unlike tissue grafts that reach a large volume 21 and 60 days after transplantation, suspension grafts do not inhibit the growth of axons of the recipient to the periphery. It was established that introduction of a suspension of dissociated cells of the spinal cord embryonic anlages (but not fragments of these anlages) into the impaired sciatic nerve in rats results in an increase in the amount of myelinated regenerating nerve fibers of the recipient 60 days after the operation.     

Opsin-immunoreactive outer segments and acetylcholinesterase-positive neurons in the pineal complex of Phoxinus phoxinus (Teleostei,Cyprinidae)

Summary The pineal complex of the teleost, Phoxinus phoxinus L., was studied light-microscopically by the use of the indirect immunocytochemical antiopsin reaction and the histochemical acetylcholinersterase (AChE) method.Opsin-immunoreactive outer segments of photoreceptor cells were demonstrated in large numbers in all divisions of the pineal end-vesicle and in the pineal stalk. Moreover, they were found in the roof of the third ventricle, adjacent to the orifice of the pineal recess as well as scattered in the parapineal organ. These immunocytochemical observations provide direct evidence of the presence of an opsin associated with a photopigment in the photosensory cells of the pineal and parapineal organs of Phoxinus. By means of the AChE reaction (Karnovsky and Roots 1964) inner segments of pineal photoreceptors, intrinsic nerve cells, several intrapineal bundles of nerve fibers, and a prominent pineal tract were specifically marked. The pineal neurons can be divided into two types: one is located near the pineal lumen, the other near the basal lamina. The latter perikarya bear stained processes directed toward the photoreceptor layer. A rostral aggregation of two types of AChE-positive nerve cells occurs in the ventral wall of the pineal end-vesicle. The main portion of the AChE-positive pineal tract, which lies within the dorsal wall of the pineal stalk, can be followed to the posterior commissure where some of the nerve fibers course laterally. A few AChE-positive pineal nerve fibers run toward the lateral habenular nucleus via the habenular commissure. In the region of the subcommissural organ single AChE-positive neurons accompany the pineal tract. The nerve cells of the parapineal organ exhibit a moderate AChE activity.These findings extend the structural basis for the remarkable light-dependent activity of the pineal organ of Phoxinus phoxinus. To the memory of Professor Karl von Frisch, pioneer and master in the field of photoneuroendocrine systemsThis investigation was supported by grants from the Deutsche Forschungsgemeinschaft to A.O. (Ok 1/24; 1/25: Mechanismen biologischer Uhren) and to H.-W. K. (Ko 758/1; 758–2)On leave from the 2nd Department of Anatomy, SOTE, Budapest, Hungary     

Origin of galanin in nerves of cat airways and colocalization with vasoactive intestinal peptide

Galanin is a 29 amino acid residue neuropeptide. In mammalian airways, galanin is found in nerve fibers associated with airway smooth muscle, bronchial glands, and blood vessels, and in nerve cell bodies of airway ganglia. The present study was conducted to determine if galanin-containing fibers in the walls of feline airways originate from the nerve cell bodies of airway ganglia. The colocalization of galanin with vasoactive intestinal peptide was also investigated. Organotypic cultures of cat airways were held in culture for 0 (nonculture control), 3, 5, and 7 days. After each culture period, the distribution of galanin and the colocalization of galanin with vasoactive intestinal peptide were determined by immunocytochemistry. Galanin-containing fibers were found in bronchial smooth muscle, around bronchial glands and in the walls of bronchial arteries and arterioles throughout the culture period. Nerve fibers and cell bodies containing both galanin and vasoactive intestinal peptide were observed after all culture periods. Nerve fibers and cells bodies that contained galanin frequently contained vasoactive intestinal peptide as well, but nerve fibers with only galanin or vasoactive intestinal were also observed. Galanin- and vasoactive intestinal peptide-containing nerve fibers and cell bodies were both well maintained throughout the culture period. The findings show that galanin-containing nerve fibers associated with bronchial smooth muscle, bronchial glands, and bronchial arteries, originate from nerve cell bodies of intrinsic airway ganglia, and that galanin and vasoactive intestinal peptide are frequently colocalized in these neurons.     

Serotonin fiber innervation of cerebellar cell suspensions intraparenchymally grafted to the cerebellum ofpcd mutant mice

One aspect of integration of implanted neurons into the neuronal circuitry of a defective host brain is the re-establishment of a host-to-graft afferent innervation. We addressed this issue by using the adult cerebellum of Purkinje cell degeneration (pcd) mutant mice, which lack virtually all Purkinje cells after postnatal day (P) 45. Purkinje cells constitute one of the cerebellar cell types being innervated by axons of raphé serotonin (5-HT) neurons. In normal mice, 5-HT-immunoreactive fibers are distributed to all cerebellar folia. Following Purkinje cell loss inpcd mice, cerebellar 5-HT-immunoreactive fibers persist. Cerebellar cell suspensions were prepared from embryonic day (E) 11–13 normal mouse embryos and were intraparenchymally grafted into the cerebellum ofpcd mutants either directly or after pre-treatment with 5, 7-dihydroxytryptamine (5,7-DHT) to selectively remove 5-HT cells of donor origin. The state of Purkinje cells and 5-HT axons was monitored in alternate sections by 28-kDa Ca²⁺-binding protein (CaBP) and 5-HT immunocytochemistry, respectively. Serotonin-immunoreactive axons were seen in the grafts from 5 to 32 days after transplantation. In some of the grafts which had not been pre-treated with 5,7-DHT, a small number of 5-HT-immunoreactive cell bodies was found, indicating that part of the 5-HT fiber innervation of the graft could actually derive from donor cells. On the other hand, in grafts pre-treated with 5,7-DHT, no 5-HT cell bodies were seen in the grafted cerebellum; 5-HT fibre innervation of the grafts occurred, but it appeared to be slightly less robust compared to situations of co-grafted 5-HT cell bodies. These findings suggest that host 5-HT fibers are able to provide afferent innervation to donor cerebellar tissue; the presence of co-grafted 5-HT cells may augment such an innervation.Special issue dedicated to Dr. Morris H. Aprison.     

Catecholamine- and acetylcholinesterase-containing nerves in human lower respiratory tract

Summary The innervation of human lower respiratory tract was studied with special emphasis on airways with sodium-potassium glyoxylic acid (SPG) and acetylcholinesterase (AChE) methods to demonstrate catecholamine-containing and acetylcholinesterase-containing nerve fibers. AChE-method revealed a rich network of cholinesterase positive nerves both inside the bronchial glands where they run around and between the acini, and the airway smooth muscle from secondary bronchi to terminal bronchioli. No AChE-positive fibers were found in connection with the blood vessels or within the epithelium of bronchi or bonchioli. The AChE-positive nerve fibers in bronchial smooth muscle greatly outnumbered those containing catecholamine. The SPG-method revealed the presence of adrenergic nerves from the level of secondary bronchi to that of terminal bronchioli. These nerve fibers were most abundant in bronchial glands, where their amount was equal and distribution similar to those of AChE-containing nerve fibers. Outside the glands adrenergic fibers were constantly seen in connection with the bronchial blood vessels in connective tissues surrounding bronchi. A few nerve fibers were also present in airway smooth muscle from the secondary bronchi to terminal bronchioli.     

The presence of acetylcholinesterase-positive nerve fibres in the deep pineal gland and the pineal stalk but not in the superficial part of adult albino rats

Application of the histochemical method for testing acetylcholinesterase (AChE, EC 3.1.1.7) showed the presence of AChE-positive nerve fibers in the deep pineal gland and the pineal stalk but not in the superficial part of adult albino rats. These findings may indirectly support the existence of the potentially cholinergic innervation of at least some of the rat pinealocytes present in these parts of the gland and augment the evidence of the heterogeneity of the rat pinealocytes. It is possible that cholinergic neurons in the medial habenular nuclei or in the parasympathetic sphenopalatine ganglion may be a source of these AChE-positive fibres. The examination was performed at the light microscope level.     

Cholinesterase activity in quail primary lymphoid organs

The present histochemical study was carried out to analyze the distribution and topography of acetylcholinepositive nerve fibers in the thymus and bursa of Fabricius of quails. The AChE-positive nerve fibers were demonstrated by direct thiocholine histochemical method. Nerve fibers present in the thymuses form periarterial nerve plexuses located mostly in the interlobular septa and on the cortico-medullary junction. Vessels-independent nerve fibers occur also in the parenchyma of thymic medulla, but rarely in parenchyma of the cortex. Within the connective tissue between the bursa of Fabricius and the wall of proctodeum we observed conspicuous AChE-positive ganglia, often in close relationship to greater arteries. Within the wall of bursa of Fabricius, AChE-positive nerve fibers create nerve plexuses around arteries. We observed a close relationship between lymphoid follicules in bursal submucosa and mucosa and AChE-positive nerve fibers. Nerve fibers create a ring around lymphoid follicles, but do not penetrate into the germinal center of the follicle. Arteries inside quail thymuses and bursae of Fabricius contain rich AChE-positive nerve plexuses, when compared to the veins, which have a very poor presence of AChE-positive nerves. According to lesser presence and decreased density of AChE-positive nerve fibers in older animals, we described age-dependent changes in both quail primary lymphoid organs.     

Vasoactive intestinal polypeptide in rat heart atria: the effect of hyperthyroidism

The effects of transient and sustained hyperthyroidism on vasoactive intestinal polypeptide-like immunoreactivity (VIP-LI) levels were studied in the heart atria of developing and adult rats. Newborn rats were divided into 5 groups. Neo-T animals were treated with thyroxine (T4) during postnatal days 1-8 and sacrificed at the age of 60 days. Neo-S rats were treated with T4 during postnatal days 1-60 and sacrificed one day later. Adult-1 and Adult-2 animals received T4 during days 52-60 and were sacrificed 5-6 days and 1 day later, respectively. Control animals were injected with saline. VIP-LI concentrations were determined in extracts from the left and right atria separately. In Neo-S and Adult-2 rats, spontaneous heart rate, the weight of both atria and total T4 serum levels were significantly enhanced, while their body weight was decreased. The ratio atria weight to body weight was significantly increased in all groups except for Adult-1 animals. Hyperthyroidism led to a significant decrease in VIP-LI levels in both atria of Neo-S and Neo-T rats. Hyperthyroidism induced in adult rats also decreased VIP-LI levels in both atria. However, this change was only transient. In conclusion, our data have provided new evidence that hyperthyroidism induced during the early neonatal period interferes with the development of VIP-ergic innervation in rat atria. The period of the first few postnatal days seems to be essential for this effect, since VIP-LI concentrations in 60-day-old animals did not significantly differ between Neo-S and Neo-T atria.     

Engraftment and differentiation of embryonic stem cell-derived neural progenitor cells in the cochlear nerve trunk: growth of processes into the organ of Corti

Hearing loss in mammals is irreversible because cochlear neurons and hair cells do not regenerate. To determine whether we could replace neurons lost to primary neuronal degeneration, we injected EYFP-expressing embryonic stem cell-derived mouse neural progenitor cells into the cochlear nerve trunk in immunosuppressed animals 1 week after destroying the cochlear nerve (spiral ganglion) cells while leaving hair cells intact by ouabain application to the round window at the base of the cochlea in gerbils. At 3 days post transplantation, small grafts were seen that expressed endogenous EYFP and could be immunolabeled for neuron-specific markers. Twelve days after transplantation, the grafts had neurons that extended processes from the nerve core toward the denervated organ of Corti. By 64-98 days, the grafts had sent out abundant processes that occupied a significant portion of the space formerly occupied by the cochlear nerve. The neurites grew in fasciculating bundles projecting through Rosenthal's canal, the former site of spiral ganglion cells, into the osseous spiral lamina and ultimately into the organ of Corti, where they contacted hair cells. Neuronal counts showed a significant increase in neuronal processes near the sensory epithelium, compared to animals that were denervated without subsequent stem cell transplantation. The regeneration of these neurons shows that neurons differentiated from stem cells have the capacity to grow to a specific target in an animal model of neuronal degeneration.     

Junctional form of acetylcholinesterase restored at nerve-free endplates.

Rat soleus muscles were ectopically innervated by implanting a foreign nerve in an endplate-free region of muscle and, 2–3 weeks later, cutting the original nerve. The junctional, 16 S form of acetylcholinesterase (AChE) and focal staining for AChE disappeared from the old endplate region within a few days after denervation. In muscles with an ectopic nerve, but not in paired control muscles, 16 S AChE and focal staining were restored in the old endplate region 1–2 weeks after denervation even though nerve fibers could not be detected in that region. These results suggest that the nerve exerts a local effect, specifying the site at which junctional AChE appears, and a nonlocal effect, perhaps mediated by muscle activity, regulating the amount of junctional AChE.     

Immunohistochemical study on the distribution of vasoactive intestinal polypeptide (VIP) containing nerve fibers in the chicken pancreas

The distribution of vasoactive intestinal polypeptide (VIP) containing nervous elements in the chicken pancreas was immunohistochemically investigated by light microscopy. Strongly VIP immunoreactive ganglia existed in the interlobular connective tissue. Ganglion containing both VIP immunoreactive and non-immunoreactive nerve cells was occasionally observed in the connective tissue. Almost all the ganglion cells also showed acetylcholinesterase (AChE) activity. No extrapancreatic nerve bundles containing VIP immunoreactive nerve fibres were detected. VIP immunoreactive nerve fibres formed plexuses in the subepithelial layer of secretory ducts and the muscle layer of small arteries. The distribution pattern of VIP immunoreactive nerve fibers was similar to that of AChE-positive nerve fibers on adjacent sections. The exocrine pancreas received a rich supply of varicose nerve fibers showing VIP immunoreactivity. B-islets also were richly innervated by VIP immunoreactive varicose nerve fibers, whereas A-islets, only poorly. These observations suggest that VIP containing nerves in the chicken pancreas have an intrinsic origin, are probably derived from VIP immunoreactive, intrapancreatic ganglion cells and innervate secretory ducts, arteries, acinar cells and B-islets, and that VIP must coexist with acetylcholine in the nervous elements.     

Fibroblast populated collagen matrix promotes islet survival and reduces the number of islets required for diabetes reversal

Islet transplantation represents a viable treatment for type 1 diabetes. However, due to loss of substantial mass of islets early after transplantation, islets from two or more donors are required to achieve insulin independence. Islet-extracellular matrix disengagement, which occurs during islet isolation process, leads to subsequent islet cell apoptosis and is an important contributing factor to early islet loss. In this study, we developed a fibroblast populated collagen matrix (FPCM) as a novel scaffold to improve islet cell viability and function post-transplantation. FPCM was developed by embedding fibroblasts within type-I collagen and used as scaffold for islet grafts. Viability and insulin secretory function of islets embedded within FPCM was evaluated in vitro and in a syngeneic murine islet transplantation model. Islets embedded within acellular matrix or naked islets were used as control. Islet cell survival and function was markedly improved particularly after embedding within FPCM. The composite scaffold significantly promoted islet isograft survival and reduced the critical islet mass required for diabetes reversal by half (from 200 to 100 islets per recipient). Fibroblast embedded within FPCM produced fibronectin and growth factors and induced islet cell proliferation. No evidence of fibroblast over-growth within composite grafts was noticed. These results confirm that FPCM significantly promotes islet viability and functionality, enhances engraftment of islet grafts and decreases the critical islet mass needed to reverse hyperglycemia. This promising finding offers a new approach to reducing the number of islet donors per recipient and improving islet transplant outcome.