An autoradiographical topological study of the epithelial cell proliferative activity in the ascending colon of the mouse using a modified technique for determining the fraction of labelled mitoses (FLM)

In a previous study of the mouse ascending colon we showed that epithelial cell proliferation is more active on the tops of mucosal folds than in the flat mucosal areas between folds. In order to define the cytokinetic characteristics of this phenomenon more precisely, we have undertaken an autoradiographic investigation with 3H-thymidine. The fraction of labelled mitoses (FLM) curves and cell labelling indices have been worked out in fold top areas (FTA) and on flat areas off the folds (OFA). A new method has been adopted to draw the FLM curves. The results show no difference between the cell cycle times or in the times corresponding to G2-phase and S-phase in FTA and OFA. In contrast, the proliferative cell compartment in FTA is 40% higher than in OFA.     

Spatial distribution and cell kinetics of the glands in the human esophageal mucosa

The glands in the human esophageal mucosa have been considered rudimentary, and have been poorly studied. Only recently, their role in the defense of the esophageal mucosa in gastroesophageal reflux disease has been put in evidence. In the present study, the presence of the different esophageal gland types was observed in 82 necropsy specimens. A precise topographical study was possible in 15 specimens. Cell proliferation parameters, namely DNA synthesis and mitotic index values in the squamous epithelium and in the esophageal glands were measured in 8 patients undergoing a surgical palliative esophagectomy, after in vivo labelling with bromodeoxyuridine. Upper mucosal, lower mucosal, and submucosal glands were observed in respectively 3.7%, 87% and 99% of the 82 esophageal specimens. Our topographical analysis showed that the upper mucosal glands, lower mucosal glands, and submucosal glands are occupying respectively 0.02%, 2.61%, and 3.96% of the esophageal surface. DNA synthesis and mitotic index values in the progenitor zone of the squamous epithelium were respectively 7.91% and 0.81%, whereas the proliferative activity in the glands was extremely low. The labelling index In the excretory ducts of the glands was only 0.07%, while no labelled cells nor mitotic figures were observed in any of the glandular acini.     

Regeneration of Vocal Fold Mucosa Using Tissue-Engineered Structures with Oral Mucosal Cells

Objectives

Scarred vocal folds result in irregular vibrations during phonation due to stiffness of the vocal fold mucosa. To date, a completely satisfactory corrective procedure has yet to be achieved. We hypothesize that a potential treatment option for this disease is to replace scarred vocal folds with organotypic mucosa. The purpose of this study is to regenerate vocal fold mucosa using a tissue-engineered structure with autologous oral mucosal cells.

Study Design

Animal experiment using eight beagles (including three controls).

Methods

A 3 mm by 3 mm specimen of canine oral mucosa was surgically excised and divided into epithelial and subepithelial tissues. Epithelial cells and fibroblasts were isolated and cultured separately. The proliferated epithelial cells were co-cultured on oriented collagen gels containing the proliferated fibroblasts for an additional two weeks. The organotypic cultured tissues were transplanted to the mucosa-deficient vocal folds. Two months after transplantation, vocal fold vibrations and morphological characteristics were observed.

Results

A tissue-engineered vocal fold mucosa, consisting of stratified epithelium and lamina propria, was successfully fabricated to closely resemble the normal layered vocal fold mucosa. Laryngeal stroboscopy revealed regular but slightly small mucosal waves at the transplanted site. Immunohistochemically, stratified epithelium expressed cytokeratin, and the distributed cells in the lamina propria expressed vimentin. Elastic Van Gieson staining revealed a decreased number of elastic fibers in the lamina propria of the transplanted site.

Conclusion

The fabricated mucosa with autologous oral mucosal cells successfully restored the vocal fold mucosa. This reconstruction technique could offer substantial clinical advantages for treating intractable diseases such as scarring of the vocal folds.     

Morphological aspects of the euphonic voice

The high quality of a euphonic voice is the result of complex interactions between many organs and systems. Vibrating vocal folds play a crucial role in this process. Their physiological motion is conditioned by the presence of the layered structure of laryngeal mucosa. In this study, we assessed the degree of dysphonia according to the Union of European Phoniatrics (UEP) scale. Videoendoscopy (VLS) and videostroboscopic (VLSS) examination of the larynx was used to visualize the vibration of the vocal folds. Morphological assessment of the inter-membranous part of the vocal fold mucosa was carried out using material collected after surgical treatment (60%) or obtained from autopsy (40%). The samples were examined by light microscopy and transmission electron microscopy. In euphonic voices, 1° of dysphonia (UEP) and the physiological endoscopic (VLS) and stroboscopic (VLSS) findings of vocal folds were registered. No morphological or ultramorphological changes were observed in the cells of the multilayered flat epithelium, basal membrane or in the stroma. Unchanged epithelial cells were situated on the basal membrane with folds. Moreover, numerous pericytes, vessels with multiplication of basal membranes, scanty collagenous fibers, plasmatic cells and lymphocytes were seen. Morphological changes with signs of atrophy and polypoid degeneration of the vocal fold mucosa were found in only 3 (15%) patients.     

Ontogeny and distribution of certain endocrine cells in the human fetal large intestine

Summary In 9 fetuses, 9 to 24 weeks-old, the occurrence and relative distribution of argentaffin cells, as well as of cells immunoreactive to somatostatin (SRIF), glucagon-like polypeptide (GLI), pancreatic polypeptide (PP) and substance P (SP) were studied in five segments of the colon (appendix, cecum, ascending colon, descending colon, and rectosigmoid). For each colonic segment, data concerned with the occurrence of endocrine cells were expressed either as mean absolute numbers of specific cells per entire mucosal section, or as cell densities per mm³ of mucosa after calculation of the mucosal volume of the sections. Argentaffin, GLI, SRIF and PP immunoreactive cells are all present in relatively large numbers, scattered along the entire length of the colonic mucosa as early as the 9th–10th week of gestation, whereas substance P-containing cells occur sporadically and first appear during the 14th–17th week. Until the 20th week, with progressing embryonic development, an increase was determined in absolute numbers per section of all types of endocrine cells in all segments of the colon. This observation is clearly related to the general growth of the colonic mucosa, since cell densities per mm³ of mucosa do not greatly change or even decrease during gestation. However, it is possible that densities of argentaffin, GLI and BPP cells increase in the appendix around the 14th–17th week of gestation. Between the 20th and 24th week, absolute numbers of cells per section remain stable or slightly increase, while cell densities tend rather to decrease in all segments. These data demonstrate that some endocrine cells are present very early in the human fetal colon, but their functional significance remains to be elucidated.This work was supported by the Institut National de la Santé et de la Recherche Médicale (INSERM)     

Non-invasive in vivo measurement of the shear modulus of human vocal fold tissue

Voice is the essential part of singing and speech communication. Voice disorders significantly affect the quality of life. The viscoelastic mechanical properties of the vocal fold mucosa determine the characteristics of the vocal folds oscillations, and thereby voice quality. In the present study, a non-invasive method was developed to determine the shear modulus of human vocal fold tissue in vivo via measurements of the mucosal wave propagation speed during phonation. Images of four human subjects' vocal folds were captured using high speed digital imaging (HSDI) and magnetic resonance imaging (MRI) for different phonation pitches, specifically fundamental frequencies between 110 and 440 Hz. The MRI images were used to obtain the morphometric dimensions of each subject's vocal folds in order to determine the pixel size in the high-speed images. The mucosal wave propagation speed was determined for each subject and at each pitch value using an automated image processing algorithm. The transverse shear modulus of the vocal fold mucosa was then calculated from a surface (Rayleigh) wave propagation dispersion equation using the measured wave speeds. It was found that the mucosal wave propagation speed and therefore the shear modulus of the vocal fold tissue were generally greater at higher pitches. The results were in good agreement with those from other studies obtained via in vitro measurements, thereby supporting the validity of the proposed measurement method. This method offers the potential for in vivo clinical assessments of vocal folds viscoelasticity from HSDI.     

The microscopic anatomy of the oesophagus, stomach and intestine of the channel catfish, Ictalurus punctatus

An anatomical study of the digestive tract of the channel catfish revealed that the oesophageal mucosa was longitudinally folded and that secondary folds were occasionally located on the primary longitudinal folds. The infoldings were more numerous near the stomach. The stratified squamous epithelium covering the folds was made up of a basal layer, large mucous cells and simple squamous cells on the surface. The epithelium on the side of the folds consisted primarily of mucous secreting cells. Taste buds were observed between mucous cells on the apical portion of the oesophageal folds and were more prevalent in the cranial part of the oesophagus. The remaining layers of the oesophagus were: a lamina propria-submucosa, tunica muscularis and adventitia or serosa.
The J-shaped stomach had two regions: a large sac-shaped region containing gastric glands and a smaller, nonglandular pyloric region. The large rugae of the stomach became gradually smaller near the pylorus. There was a well developed pyloric sphincter. The mucosa included a simple columnar epithelium, a lamina propria and adventitia or serosa.
The intestine could be differentiated into a thick ascending segment, a descending segment, a thin convoluted segment and a thicker terminal segment, the rectum. Many mucosal folds containing branched villi characterized the ascending segment of the intestine. The descending and convoluted segments contained fewer folds with shorter and less-branching villi and were smaller in diameter and thinner walled. Descending and convoluted segments were also mildly convoluted and accounted for 80% of the total length of the intestine. An intestinal valve with a sphincter marked the beginning of the rectum. There was an approximately four-fold increase in the thickness of the tunica muscularis of the terminal segment of the intestine.     

The role of the mesenchyme in mouse neural fold elevation. I. Patterns of mesenchymal cell distribution and proliferation in embryos developing in vitro

Using the computer-assisted method of smoothed spatial averaging, spatial and temporal patterns of cell distribution and mitotic activity were analyzed in the cranial mesenchyme underlying the mesencephalic neural folds of mouse embryos maintained in roller tube culture. Total cell density increased in central and medial mesenchymal regions after 12 hr in culture, decreased after 18 hr, and showed a further decrease after 24 hr when the neural folds of the embryos had elevated, converged, and were fusing or fused. Mitotic activity, as measured by the ratio of 3H-thymidine-labeled cells to unlabeled cells, was highest in the central mesenchyme at all culture times. Embryos were also cultured in the presence of diazo-oxo-norleucine (DON), which inhibits glycosaminoglycan and glycoprotein synthesis. After 24 hr in culture, neural folds of DON-treated embryos had failed to elevate. Total cell density increased in central and medial regions of the mesenchyme of DON-treated folds at 12 hr but showed no significant decrease in these regions with further culture. Mitotic activity was highest in the central mesenchyme of these treated embryos. These results suggest that cell distribution patterns observed in the cranial mesenchyme during neural fold elevation in normal cultured embryos are not produced by regional differences in mitotic activity. Rather, we propose that cell distribution patterns in the central and medial regions of the mesenchyme result from expansion of a glycosaminoglycan-rich extracellular matrix that disperses cells from these regions and decreases their density. In DON-treated embryos, in which expansion of the mesenchyme is prohibited by the decreased glycosaminoglycan and glycoprotein content of the extracellular matrix, mitotic activity apparently determines these patterns.     

Comparative microscopical study of the gall bladder mucosa.

The gall bladder from 6 Psammophis sibilans, 10 Bufo regularis and 10 Albino mice were extracted and prepared for microscopic examination. It was found that the mucosa of Psammophis sibilans consisted of ovoid and polygonal cells which were occasionally binucleated cells with darkly stained nuclei and occasionally pear-shaped cells with vesicular nuclei and fine processes. These cells were arranged in three layers. Apossible explanation for the different types of cells encountered and their arrangement was given. The gall bladder mucosa of Bufo regularis and Albino mouse were thrown into folds covered with simple columnar epithelium. However, the epithelium of the frog was higher than that of the mouse, with the nuclei situated midway between basement membrane and the lumen. Vacuolated cells were detected in the gall bladder mucosa of the mouse. The significance of the mucosal folds was discussed.     

The effect of single and of multiple doses of prednisolone tertiary butyl acetate on cell population kinetics in the small bowel mucosa of the rat.

The effect of single and of multiple doses of prednisolone upon cell population kinetics in the rat jejunal crypt was investigated, using autoradiography and stathmokinetic techniques with vincristine. Single injections of prednisolone (2.5 mg/kg body weight) induced a depression both flash thymidine labelling and mitotic indices; this change was shown to be due to a decreased cell production rate. Recovery of these proliferative indices occurred over seven days after injection; measurement of crypt size parameters showed a transient decrease in crypt population. Multiple daily injections of prednisolone (1 mg/kg body weight) produced a more sustained decrease in labelling and mitotic indices, which lasted as long as injections were continued (7 days); stathmokinetic techniques showed decreases in cell production rates, and the crypt population was also depressed throughout this period. It is concluded that prednisolone depresses cell proliferative rates in rat jejunal mucosa.     

FOXP3+ cell density in lymphoid follicles from histologically normal mucosa is a strong prognostic factor in early stage colon cancer

There are few clearly established prognostic factors available to guide the use of adjuvant chemotherapy in early stage colon cancer patients. Some of the most promising candidates include the invasion of extramural blood vessels by tumour cells and the densities of FOXP3+ T regulatory cells (Tregs) in tumour and adjacent normal colonic mucosal tissue. The aim of our study was to evaluate the prognostic significance of these markers in AJCC stage II colon cancer, with particular reference to lymphoid follicles in the mucosa. Histopathological review for the presence of vascular and serosal invasion was conducted on a series of 165 stage II colon cancers treated by surgery alone. Immunohistochemical staining for FOXP3 was performed on tumour tissue and histologically normal colonic mucosa from the surgical margin. Image analysis software was used to evaluate the density of FOXP3+ cells in the tumour core, invading margin and lymphoid follicles from the colonic mucosa. For survival analysis, cases were classified into high- or low-density of FOXP3+ cells according to the median value. The mean density of FOXP3+ Tregs in lymphoid follicles was twofold and fivefold higher than in the invading margin and tumour core, respectively. Multivariate analysis identified extramural vascular invasion (HR, 2.47; 95% CI: 1.00-6.07; P?=?0.05) and high FOXP3+ cell density in lymphoid follicles (HR, 4.22; 95% CI: 1.49-11.91; P?=?0.007) as independent factors for worse survival, whereas a high frequency of lymphoid follicles in histologically normal colonic mucosa was associated with better survival (HR, 0.31; 95% CI: 0.12-0.79; P?=?0.014). Our data suggest that host factors related to the immune system have major prognostic significance in early stage colon cancer. The density of FOXP3+ cells within lymphoid follicles and the frequency of these structures in normal colonic mucosa represent novel and independent prognostic factors.     

Effect of ultrasound-facilitated fixation on oral mucosa density and morphology

We investigated the effects of ultrasound-facilitated fixation on oral mucosal morphology. Bovine dorsal tongue and porcine buccal (cheek) mucosa were sonicated for 0, 5, 10 or 15 min in a modified methacarn fixative, then incubated at 25° C for 4 h. Initial mass, volume and density of each specimen were measured before and after treatment and fixation. Morphometric analysis of the scanning electron micrographs was used to quantify changes in mucosal structure and microtexture. Statistical methods were used to describe the relation between sonication time, tissue density and relative change in tissue density. Our results indicate a linear correlation between sonication time and density of the dorsal tongue specimens. The treatment caused contraction of the tongue tissue and expansion of the buccal mucosa. Differences between initial and final tongue densities and the relative change in tissue density of the tongue vs. buccal mucosa were statistically significant. Differences observed between initial and final buccal mucosa densities were not statistically significant. Changes in buccal mucosa density correlated inversely with sonication time by contrast to the tongue density, which was correlated directly with this factor. Our study illustrates that while preservation of mucosal morphology and biopolymers can be achieved by ultrasound-facilitated fixation, its effects on tissue density are both time-dependent and specific to certain regions of the mouth.     

PROLIFERATION KINETICS IN EPITHELIUM OF GUINEA-PIG COLON

Autoradiographs of crypts of guinea-pig colon labelled with ³H-thymidine were quantitatively analysed. The analysis involved: (a) comparison of proliferation parameters in crypts situated in two well-defined regions of colonic folds, i.e. in the top and the basal parts of the fold, and (b) comparison of proliferation parameters in the lower and in the upper parts of an individual crypt. The results provide evidence that: (1) Crypts situated in the top region of folds are significantly longer than those situated in the basal region. (2) The duration of cell cycle and its phases is similar in the crypts of the two regions examined. (3) The rate of ³H-thymidine uptake is higher in the middle than in the initial and final parts of the S phase, and its pattern is the same for the crypts of the two regions of the fold as well as for the parts of the individual crypt. (4) The proliferating pool size is larger in crypts situated in the top than in the basal region of the fold. (5) The proliferating pool size is larger in the lower parts of crypts from each region than in the upper parts. Control of cellular proliferation in the crypt epithelium is considered to operate separately at the level of the individual crypt and at the level of a group of crypts situated in a given region of the colonic wall.     

Effect of starvation on endocrine cells in the rat stomach

The influence of food deprivation on gastric G- and D-cells and on parietal cells was studied in the rat. In fed controls and groups of rats fasted for 12 and 96 h G-, D- and parietal cell densities, somatostatin and gastrin concentration in antral and fundic specimens and serum gastrin were compared. Gastrin in antral mucosa, serum gastrin, G-cell density as well as antral D-cell density decreased in long-term fasted rats by 52%, 90%, 58% and 42%, respectively. Fundic D-cell density remained unchanged. After 96 h starvation somatostatin concentration slightly increased in antral mucosa (+35%; P less than 0.05), but decreased in fundic mucosa (-40%; P less than 0.05). Parietal cell density was not influenced by prolonged fasting. These findings demonstrate that changes in D-cell morphology and mucosal somatostatin content are not parallel and that the rat gastric D-cell is less dependent on food in the gastric lumen than the G-cell. The unaltered fundic D-cell density reflects the functional activity of gastric D-cell which has also been shown to be independent of the presence or absence of food.     

Endometrial vascularity during pseudopregnancy in the rabbit

The vascular architecture in the rabbit uterus was studied during pseudopregnancy. Uteri at 2, 3, 4, 6, 7, 8, 9, 11, 13, 16, 17, 18, 20, 24 and 28 days after sterile mating were subjected to one of four techniques: neoprene latex casts, transparent sections, frozen sections, and histological sections. Measurements were made microscopically of the thickness of myometrium and of the subepithelial capillary plexus in the different mucosal folds. In the estrous rabbit, the circular arteries in the uterine muscular layer give off arterioles which pass upwards, with a few branchings through the endometrium to the uterine lumen. These arterioles reach the surface of the mucosal folds and break up into the subepithelial capillary plexus. This plexus is connected to the tips of the venules which run down through the endometrium to the endometrial vascular plexus at the base of the endometrium: some of the venules connect with the circular venous vessels in the muscular layer. With advanced stages of pseudopregnancy, the capillaries among the glands become stretched and elongated. Maximal branching of the folds occurs at 4 to 9 days of pseudopregnancy. The “branching activity” was consistently higher in the placental than in the periplacental or in the obplacental folds. Such changes reached a maximum at 6 to 7 days p.c., after which the capillaries became gradually shorter and tortuous. The development of arterioles in the mucosa was marked at 3 to 6 days p.c. The thickness of the plexus in the periplacental fold and in the obplacental fold as a percentage of the thickness in the placental fold was highly correlated with the stage of pseudopregnancy. At 9 to 11 days p.c., these ratios reached a minimum of 70 to 80%.     

Possible relationship between Helicobacter pylori infection and cap polyposis of the colon

BACKGROUND: Cap polyposis is a rarely encountered disease characterized by multiple distinctive inflammatory colonic polyps located from the rectum to the distal colon. The etiology of this disease is still unknown, and no specific treatment has been established. AIM: We report three cases of cap polyposis that were cured following eradication therapy for Helicobacter pylori infection. METHODS AND RESULTS: Three women were referred to Shinshu University Hospital because of mucoid and/or bloody diarrhea. Laboratory data showed hypoproteinemia in all cases; markers of inflammation such as C-reactive protein were negative. Colonoscopy revealed multiple sessile polyps with mucus adherent on the apices of the mucosal folds in the rectum and/or the sigmoid colon. The intervening mucosa was normal. Microscopic examinations of biopsy specimens taken from sessile polyps revealed inflamed mucosa with elongated tortuous crypts attenuated towards the mucosal surface. A granulation tissue 'cap' was observed on the surface of the mucosa. Various treatments were unsuccessful, including administration of metronidazole or prednisolone, avoidance of straining at defecation, and surgical or endoscopic resection. All were diagnosed with H. pylori infection in the stomach. Helicobacter pylori was not detected in the biopsy specimens from the colonic inflammatory polyps by immunohistochemical study using polyclonal anti-H. pylori antibody. After successful eradication therapy the clinical symptoms improved. Disappearance of cap polyposis was confirmed by colonoscopy in all three cases. CONCLUSION: We speculate that H. pylori infection might play a role in the pathogenesis of cap polyposis.     

泽陆蛙消化道组织学观察

采用解剖学和组织学的方法对泽陆蛙消化道各部分的形态和组织学结构进行了观察。泽陆蛙的消化道分为口腔、咽、食道、胃、十二指肠、回肠和大肠。各管壁都由黏膜层、黏膜下层、肌层和外膜组成。食道黏膜层有皱褶和纤毛,无杯状细胞;胃黏膜层有杯状细胞和胃腺,黏膜下层有血管分布;小肠具绒毛、杯状细胞和肠腺,大肠皱褶少,杯状细胞也少。     

Melatonin-induced suppression of the pinealectomy-stimulated rat adrenal cortex mitotic activity

Pineal involvement in the regulation of adrenocortical mitotic activity has recently been suggested. It has been shown that melatonin (Mel) decreased the mean mitotic activity rate (MMAR) of the adrenal cortex both in vivo and in organ culture. The goal of the present study was to test the influence of pinealectomy (PX) and/or Mel-treatment on the MMAR of adrenocortical cells, as well as on the adrenal weight in rats. The stathmokinetic method was used in the study. It was found that PX significantly increased the MMAR of the adrenocortical cells. Moreover, Mel suppressed the proliferogenic effect of PX on the rat adrenocortical cells. Melatonin alone did not significantly affect the mitotic activity of the adrenal cortex. None of the three experimental procedures, i.e. Mel, PX and Mel-treatment of pinealectomized animals significantly affected the adrenal weight. The present data suggest that Mel may be involved in the inhibitory control of adrenocortical cell proliferation.     

Effects of diazepam on cell proliferation in cerebral cortex, anterior pituitary and thymus of developing rats

The effect of a single dose (5mg/kg b.w.) of diazepam on the cell proliferation in selected organs of 11-day old female Sprague-Dawley rats has been investigated. The stathmokinetic method (counting of mitoses after colchicine administration) has been used for evaluation of cell replication. The significant fall of the mitotic activity in the parietal cerebral cortex and the anterior pituitary gland was found. On the other hand, the increase of the mitotic activity in thymus was observed. The reported data, taken together with the previous observations from our and other laboratories, suggest the involvement of benzodiazepine receptor in the control of cell proliferation.     

Cell population kinetics in the epithelium of the colon of the male rat.

The present study was undertaken in order to try to define some of the kinetic parameters in the colonic mucosa of normal Wistar rats. Preliminary observations showed considerable morphological differences in the mucosa from site to site along the length of the colon. In particular the height of the crypts (measured in cells) was variable. In addition labelling index studies demonstrated dramatic variations in the distribution of labelling along the length of the crypts from site to site in the bowel. A single site in the descending colon was selected for more detailed study using a stathmokinetic agent, vincristine, and the continous labelling technique with tritiated thymidine. The results of these investigations suggest that there exists at the base of the crypt a subpopulation of cells cycling more slowly than the cells in the rest of the proliferative compartment. Growth fraction appears to fall with rising cell positions within the crypt.