Gian-Luca Ferri

Neuron Specific Enolase: A Common Marker for the Endocrine Cells and Innervation of the Gut and Pancreas

Neuron specific enolase, the most acidic isoenzyme of the glycolytic enzyme enolase, was first believed to be present exclusively in central neurons. More recently, it has been found in peripheral autonomic nerves and in a number of endocrine cells. An immunocytochemical study was carried out concerning the distribution of neuron specific enolase in the gastrointestinal tract and pancreas of humans and rats. In addition, immunocytochemistry and histochemistry were used to obtain a characterization of the different types of cells and nerves in which neuron specific enolase can be detected. Neuron specific enolase was found in all currently identifiable endocrine cell types and nerves of the gut and pancreas. Neuron specific enolase is therefore a common marker for both endocrine cells and enteric nerves, thus providing a simple means for their simultaneous demonstration and examination of their morphologic characteristics and integration.

Neuronal intermediate filaments in rat dorsal root ganglia: differential distribution of peripherin and neurofilament protein immunoreactivity and effect of capsaicin.

Two major neuronal populations were revealed in rat dorsal root ganglia, immunoreactive for either peripherin, or neurofilament triplet proteins (adult L2 ganglia: 66.2% and 25.6%, mainly small and large diameter cells, respectively), together with a minor, double-immunostained population (L2: 8.1%, mainly intermediate-size neurons). After capsaicin treatment, a striking expansion in the latter population was seen (L2: 22.0%) together with a significant increase in size, restricted to the same population and the (remaining) peripherin-only immunoreactive neurons. Calcitonin gene-related peptide (CGRP) immunoreactivity was revealed in neurons of all 3 groups, in both normal and capsaicin-treated rats.

Intramural distribution of immunoreactive vasoactive intestinal polypeptide (VIP), substance P, somatostatin and mammalian bombesin in the oesophago-gastro-pyloric region of the human gut

SummaryThe intramural distribution of vasoactive intestinal polypeptide (VIP), substance P, somatostatin and mammalian bombesin was studied in the oesophago-gastro-pyloric region of the human gut. At each of 21 sampling sites encompassing this entire area, the gut wall was separated into mucosa, submucosa and muscularis externa, and extracted for radioimmunoassay. VIP levels in the mucosa were very high in the proximal oesophagus (1231±174 pmol/g, mean±SEM) and showed varied, but generally decreasing concentrations towards the stomach, followed by a clear-cut increase across the pyloric canal (distal antrum: 73±16 pmol/g, proximal duodenum: 366±62 pmol/ g); consistent levels were found in submucosa and muscle (200–400 pmol/g) at most sites, the stomach again showing lower concentrations. By contrast, substance P was present in small amounts as far as the proximal stomach, but sharply increased across the pyloric canal, especially in mucosa and submucosa (distal antrum: 20±6.5 and 5.5±1.3 pmol/g; proximal duodenum: 62±8.5 and 34±11 pmol/g, respectively). Somatostatin concentrations were very low in the mucosa of the oesophagus and stepwise increased in the cardiac, mid-gastric and pyloric mucosa (cardia: 224±72 pmol/g; distal antrum: 513±152 pmol/g; proximal duodenum: 1013±113 pmol/g); concentrations in the submucosa and muscularis were generally low, with the exception of antrum and duodenum. Mammalian bombesin was comparatively well represented throughout the oesophageal muscularis (5–8 pmol/g), but most abundant in the stomach in all layers (oxyntic mucosa: 24±2.7 pmol/g; submucosa: 20±5.7 pmol/g; muscle: 28±5.0 pmol/g). In conclusion, a distinct differential distribution of the four peptides studied was revealed, indicating a diffuse, but highly differentiated peptide-containing innervation of the proximal human gut.

Heal Enteroglucagon Cells after Ileal-Duodenal Transposition in the Rat

The changes occurring in the ileal wall and in enteroglucagon cells were studied in a rat model of intestinal adaptation, obtained by the transposition of a segment of distal ileum into the mid-duodenum (6 rats, compared with 6 transected controls). After 40 days, the transposed ileal segment, compared to the equivalent segment in the controls, showed striking increase in weight, especially of the epithelium (1,585 +/- 127 vs. 305 +/- 42 mg, mean +/- SEM, p less than 0.0005). The calculated weight of enteroglucagon cells in the segment showed a smaller, but significant increase (1.7 +/- 0.3 vs. 0.8 +/- 0.2 micrograms, p less than 0.05). Plasma enteroglucagon was markedly raised (239 +/- 28 vs. 61 +/- 7.1 pmol/l, p less than 0.0005) and showed a greatly increased meal-stimulated response (1,521 +/- 284 vs. 83 +/- 43 pmol, p less than 0.0005), thus suggesting hyperactivity of enteroglucagon cells.

Immunocytochemical evidence for the presence of γ1-MSH-like immunoreactivity in pituitary corticotrophs and ACTH-producing tumours

The presence of gamma 1-MSH has been demonstrated in bovine neuro-intermediate lobe by biochemical methods, thus suggesting that this peptide is cleaved from the cryptic region of pro-opiocortin. In this study we report the localisation of gamma 1-MSH-like immunoreactivity in the adenohypophysis of man, ox, pig, dog and guinea-pig using immunocytochemical procedures at both light and electron microscope levels. Antisera recognising the C-terminal Arg-Phe-amide and the C-terminal penta-peptide-amide of gamma 1-MSH have been used throughout this study. The immunostaining was found in all endocrine cells of the pars intermedia (where present) and in scattered cells of the pars distalis identified as corticotrophs. No gamma 1-MSH immunoreactivity was detected in rat adenohypophysis. In addition, 7 ACTH-producing tumours (1 pituitary adenoma and 6 ectopic) were investigated and shown to contain gamma 1-MSH immunoreactive cells.

Intramural distribution of Met5-enkephalin-Arg6-Gly7-Leu8 in sphincter regions of the human gut

The intramural distribution of Met5-enkephalin-Arg6-Gly7-Leu8 (MERGL) was studied in the oesophago-cardiac, pyloric, ileo-caecal and sigmoid-recto-anal regions of the human digestive tract. Serial samples encompassing each area were separated into mucosa, submucosa and muscularis externa and extracted for radioimmunoassay. Comparatively low levels of MERGL immunoreactivity were measured throughout the cardiac junction. Conversely, a remarkable peak of MERGL concentration was detected at the pyloric junction, in both submucosa and muscularis. A progressive decrease in tissue levels of the same peptide, most evident in the submucosa, was detected on the proximal side of the ileo-caecal region. In the distal sigmoid colon and rectum MERGL concentrations showed a rapid decline, down to very low levels in the anal canal. The results may suggest the involvement of an enkephalinergic mechanism in the control of the human pylorus.

Aluminum Foil Molds for Cryostat Blocks

Inexpensive pyramidal templates may be prepared in a range of sizes from segments of plastic or wooden bars (Fig. 1). Pieces of double-layered aluminum cooking foil are shaped by hand around templates of appropriate size and their surfaces are smoothed with fingernails. In this way, “negative” molds of known cross-section and variable depth can be obtained (Fig. 1). Tissue samples are oriented on the bottom of the molds in a few drops of Ames O.C.T. Compound and just covered with the same embedding medium. Narrow strips of light-weight cardboard with identification data are inserted into the blocks close to a lateral surface. The blocks are solidified on the cold stage of a cryostat (a drop of O.C.T. will improve thermal contact and help keep them upright), or with a liquid gas, such as nitrogen, or Freon 22 (in the latter case the molds are immersed as far as the upper level of the O.C.T., but are not submerged, so that the Freon does not bubble into the liquid O.C.T.). After freezing, the foil is remove...

2,5-Hexanedione-induced accumulations of neurofilament-immunoreactive material throughout the rat autonomic nervous system.

In rats intoxicated with 2,5-hexanedione, nerve fibres supplying virtually all visceral organs showed large numbers of densely immunoreactive accumulations of neurofilament-like material, of fusiform, elongated, smoothly tapering morphology. In the gut, round to oval, morphologically different lesions were also present, and abnormal neurofilament-immunoreactive accumulations were revealed in oesophageal terminal end-plates. An extensive damage to autonomic nerve fibres, which are largely non-myelinated, was thus revealed in 2,5-hexanedione intoxication. The observed diversity in lesion morphology may suggest heterogeneity in cytoskeletal and/or associated proteins among autonomic neurons.

Regulatory peptide distribution in separated layers of the human jejunum.

The distribution of regulatory peptides was studied in the separated epithelium, lamina propria, submucosa and muscularis externa of the human jejunum. Gastrin, secretin, gastric inhibitory polypeptide, enteroglucagon and neurotensin immunoreactivity were almost confined to the endocrine cell-containing mucosal epithelium (greater than 98% of the total content), only minor amounts of motilin being detected in non-epithelial layers (3.6 +/- 0.7%, mean +/- SEM, n = 7). Conversely, vasoactive intestinal polypeptide, substance P and mammalian bombesin were virtually limited to non-epithelial layers (greater than 99%). Only somatostatin was found in all layers (44 +/- 6.7% in the epithelium, 34 +/- 5.2% in the lamina propria, 13 +/- 2.9% in the submucosa, and 7.9 +/- 2.8% in the muscularis). Substance P was found in higher concentrations in the mucosa, compared to submucosa and muscle (56 +/- 10, 30 +/- 4.0 and 29 +/- 4.0 pmol/g, respectively), while vasoactive intestinal polypeptide was more abundant in the muscle (411 +/- 52 pmol/g) compared to mucosa and submucosa (228 +/- 64 and 219 +/- 31 pmol/g, respectively). Only low levels of mammalian bombesin were measured, mainly in the muscle (6.9 +/- 1.5 pmol/g, or 89 +/- 3.6% of total content).