phytobenzoars-like structure covers the mucosal surface of...

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OPEN ACCESS Asian Journal of Animal and Veterinary Advances

ISSN 1683-9919DOI: 10.3923/ajava.2016.709.716

Research ArticlePhytobenzoars-like Structure Covers the Mucosal Surface ofSunda Porcupines (Hystrix javanica) Stomach1Teguh Budipitojo, 2Yuda Heru Fibrianto and 3Guntari Titik Mulyani

1Department of Anatomy,2Department of Physiology,3Department of Internal Medicine, Faculty of Veterinary Medicine, Gadjah Mada University, 55281 Yogyakarta, Indonesia

AbstractBackground and Objective: The present study was conducted to determine the histological structure of mucosa stomach in adult sundaporcupine (Hystrix javanica) which is endemic to Indonesia. Sunda porcupine is an important vertebrate pest in Indonesia’s agriculture,but there is little knowledge of stomach structure of this pest. Materials and Methods: During the present study, five Hystrix javanicawere used to examine the structure of mucosa stomach, microscopically by using hematoxylin-eosin and alcian blue-periodic acid Schiff’sstaining methods. Results: Histologically, the wall of the tunica mucosa stomach consists of three laminas, lamina epithelialis mucosa,lamina propria mucosa and lamina muscularis mucosa. The stomach covered by thick phytobenzoars-like structure which fused withsimple short columnar epithelium that extends through gastric pits, where the gastric glands are opened. Three regions can be recognizedin the mucosa of the stomach according to the types of glands that appear as tubular short glands with bent portion at the basal in cardiacregion, tubular branched and coiled glands in fundic glands region while, the pyloric glands region appear as simple branched tubularlong straight over most of their length parallel to each other. The cardiac glands noted mainly serous secreting cells while the fundicglands region consist of different types of cells but the abundance main cells is the parietal cells. Pyloric glands region consist of two maincells are the parietal and chief cells, but the number of parietal cells dramatically decrease compare with fundic glands region andgradually dissapear in the end of the region. Conclusion: The tunica mucosa stomach of Hystrix javanica consists of three lamina (laminaepithelialis mucosa, lamina propria mucosa and lamina muscularis mucosa) and divided into three gland regions (richly neutralmucopolysacarides producing cells of cardiac, acidic mucopolysacarides producing cells of fundic, mixed acid and neutralmucopolysacarides producing cells of pyloric glands regions) covered by neutral mucopolysacarides of thick phytobenzoars-likestructures.

Key words: Herbivore rodent, Hystrix javanica, stomach, tunica mucosa, cardiac gland region, fundic gland region, pyloric gland region, phytobenzoars-likestructure, HE, AB-PAS

Received: April 25, 2016 Accepted: September 05, 2016 Published: October 15, 2016

Citation: Teguh Budipitojo, Yuda Heru Fibrianto and Guntari Titik Mulyani, 2016. Phytobenzoars-like structure covers the mucosal surface of sundaporcupines (Hystrix javanica) stomach. Asian J. Anim. Vet. Adv., 11: 709-716.

Corresponding Author: Teguh Budipitojo, Department of Anatomy, Faculty of Veterinary Medicine, Gadjah Mada University, 55281 Yogyakarta, Indonesia

Copyright: © 2016 Teguh Budipitojo et al. This is an open access article distributed under the terms of the creative commons attribution License, whichpermits unrestricted use, distribution and reproduction in any medium, provided the original author and source are credited.

Competing Interest: The authors have declared that no competing interest exists.

Data Availability: All relevant data are within the paper and its supporting information files.

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Asian J. Anim. Vet. Adv., 11 (11): 709-716, 2016

INTRODUCTION

The vertebrate alimentary canal is a highly specializedstructure that brings food into an organism, digests the food,absorbs nutrients and expels waste products1. Digestive tractmorphology can affect digestive efficiency and is closelyrelated to food habits. Feeding habits and ecologicalconditions lead to changes in the basic metabolic rate whichresult in variation in dimensions of the alimentary canal2-4.Within different groups of animals, the metabolic rate isconnected with the type of a diet, structure and function ofthe alimentary canal5,6.

Differences in food habits among mammals are oftenreflected in the structure of the alimentary tract7. Vorontsov8

has observed differences in digestive tract morphology toevolutionary adaptations for a herbivorous diet. The transitionfrom a high-energy, high-protein and high-lipid diet of seedsand small invertebrate, to a low-energy and high-cellulose dietof vegetative parts of plants was hypothesized to haveresulted in several evolutionary modifications in the digestivetracts of rodents.

The porcupine belongs to the Hystricidae family whichconstitutes a small group of the order Rodentia8. Hystrixjavanica or sunda porcupine that belongs to Acanthionsubgenus of Hystricidae family is herbivorous porcupinespecies endemic to Indonesia9. There are no data related themorphological structure of stomach porcupine at present.This study was aimed to describe the mucosal stomach ofsunda porcupine, as ones of Indonesian endemic porcupines,histologically to clarify their morphological features andmucopolysacarides contents.

MATERIALS AND METHODS

Five adult (three males and two females) sundaporcupines (Hystrix javanica) were studied. Deep anesthesiaof animals was induced by initial injection of cetanes (ketamin HCL, 10-15 mg kgG1, intra muscular) followed byrompun (xylazine HCL, 0.10-0.15 mg kgG1, intra muscular)before taking the samples of stomach tissues.

The stomach tissue samples were fixed in Bouin’ssolution, dehydrated in graded series of ethanol and finallyembedded in paraffin wax. The sections were cut at 5 :m inthickness using a rotary microtome, deparaffinized andstained with Haematoxylin-Eosin (HE) method to confirm thehistological features and alcian blue in combination withperiodic acid Schiff’s (AB-PAS) technique for detection of

neutral and acid mucins and then examined descriptivelyunder a light microscope. For AB-PAS staining, deparaffinizedtissue sections were brought to distilled water throughdowngraded ethanol series and immersed in 1% AB (Sigma)in 3% acetic acid (pH 2.5) for 45 min. After washing in distilledwater, the sections were oxidized in 0 and 5% aqueousperiodic acid solution for 10-15 min at room temperature. Thesections were then immersed in Schiff’s reagent for 45 minand washed again in water. Then the slides were finallywashed in running tap water, dehydrated through ascendingseries of ethanol, cleared in xylene and mounted.

RESULTS

This study showed the histological structure of tunicamucosa of sunda porcupine stomach without sexualdistinction. The mucosa of stomach in sunda porcupineappear as divided into three regions according to the types ofglands which is contains, the very narrow cardiac, large fundicand pyloric glands regions (Fig. 1). The tunica mucosa ofstomach in cardiac, fundic and pylorus gland regions has thesame three lamina, lamina epithelialis mucosa, lamina propriamucosa and lamina muscularis mucosa. The entire surface ofgastric mucosa was lined by a simple short columnarepithelium which form the surface mucous lining cells thatinvaginate into varying depth into the lamina propriaaccording to the regions of stomach in sunda porcupineforming the gastric pits that lined by the same surfaceepithelium and where the glands are opened in the base of it(Fig. 1, 2). In all regions, the lamina epithelialis mucosa coveredby thick phytobenzoars-like structure which fused with shortcolumnar epithelium that extends through gastric pits wherethe gastric glands are opened (Fig. 1, 2).

The regions of the mucosa stomach can be easilyrecognized according to the types of glands that appear astubular short glands with bent portion at the basal in cardiacregion, tubular branched and coiled glands in fundic glandsregion while, the pyloric glands region appear as simplebranched tubular long straight over most of their lengthparallel to each other (Fig. 1, 2).

The cardiac glands noted mainly serous secreting cellswhile the fundic glands region consist of different types ofcells but the abundance main cells is the parietal cells(Fig. 1, 2). Pyloric glands region consist of two main cells arethe parietal and chief cells, but the number of parietal cellsdramatically decrease compare with fundic glands region andgradually disappear in the end of the region (Fig. 1, 2).

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Fig. 1(a-c): Histological structure of tunica mucosa of sunda porcupine stomach (HE, 130x), the stomach mucosa of sundaporcupine divided into three regions according to the types of glands which is contains, (a) Very narrow (black b)cardiac, (b) Large fundic, (c) Pyloric glands regions, all of the regions have the same three lamina, lamina epithelialismucosa (white c), lamina propria mucosa (white b), lamina muscularis mucosa (white a) and the lamina epithelialismucosa covered by thick phytobenzoars-like structure which fused with short columnar epithelium (white c)

The Alcian Blue-Periodic Acid Schiff’s (AB-PAS) stain hasbeen used to high light the contents of phytobenzoars-likestructures and mucus-secreting cells of the gastric mucosa.

The surface lining cells of entire mucosa of sunda porcupinestomach covered by thick phytobenzoars-like structure thatgave a strong reactivity with PAS which appear as pink color

711

(a) (b)

(c)


Fig. 2(a-i): Histological structure of tunica mucosa of sunda porcupine stomach (HE, 520x), (a) In the cardiac, (b) Fundic, (c) Pyloricglands regions, the lamina epithelialis mucosa covered by thick phytobenzoars-like structure (black lines) which fusedwith short columnar epithelium (e) that extends through gastric pits (stars) where the gastric glands are opened,(d) The regions of the mucosa stomach can be easily recognized according to the types of glands that appear astubular short glands middle part, (e) In cardiac region, tubular branched and coiled glands in the middle, (f) While themiddle (g) With bent portion at the basal part, (h) Basal parts of fundic glands region and (i) Middle parts of pyloricglands region appear as simple branched tubular long straight over most of their length parallel to each other, thecardiac glands noted mainly serous secreting cells (thin arrows) while, the fundic glands region consist of differenttypes of cells but the abundance main cells is the parietal cells (thick arrows), pyloric glands region consist of two maincells are the parietal (thick arrows) and chief (thin arrows) cells but the number of parietal cells dramatically decreasecompare with fundic glands region and gradually dissapear in the end of the region

and considered as neutral mucopolysacarides (Fig. 3). At themiddle and basal parts of the gastric mucosa, the seroussecreting cells of the cardiac glands region are gave a positivereaction with PAS and appear as a magenta color andconsidered as neutral mucopolysacarides and fundic glands

region are gave a positive reaction with AB and appear as ablue color and considered as acid mucopolysacarides andpyloric glands region are gave a positive reaction withAB-PAS and appear as blue and pink colors,considered as acidand neutral mucopolysacarides (Fig. 3).

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(a)

(d)

(g)

(b) (c)

(e) (f)

(h) (i)


Fig. 3(a-i): The mucopolysacarides products and cell types of mucosa stomach of sunda porcupine (AB-PAS, 520x), (a) The surfacelining cells of gastric, (b) Fundic, (c) Pyloric mucosa of sunda porcupine stomach covered by thick phytobenzoars-likestructure that gave a strong reactivity with PAS which appear as pink color and considered as neutralmucopolysacarides which indicated by arrows, (d, g)At the middle and basal parts of the gastric mucosa respectively,the serous secreting cells of the cardiac glands region are gave a positive reaction with PAS and appear as a magentacolor and considered as neutral mucopolysacarides which indicated by white stars, (e, h)Abundance parietal cells ofthe fundic glands region are gave a positive reaction with AB and appear as a blue color and considered as acidmucopolysacarides which indicated by black stars and (f, i) While in the pyloric glands region a small number of parietalcells are gave a positive reaction with PAS and appear as pink colors as neutral mucopolysacarides indicator (whitestars)

713

(a) (b) (c)

(d) (e) (f)

(g) (h) (i)


DISCUSSION

The present study revealed that the mucosa of stomachin sunda porcupine appear as divided into three regionsaccording to the types of glands which is contains the verynarrow cardiac and large area of fundus and pyloric glandsregions. The general wall structure of tunica mucosa stomachin cardiac, fundic and pylorus gland regions of sundaporcupine which composed of three lamina, laminaepithelialis mucosa, lamina propria mucosa and laminamuscularis mucosa is similar to that previously described inmonogastric animals reported by Ghoshal and Bal10 and insome mammalian stomach11.

Differ from the other stomach of both ruminant12 andnon-ruminant13 in normal condition, the lamina epithelialismucosa of normal sunda porcupine covered by thickphytobenzoars-like structure which fused with short columnarepithelium that extends through gastric pits where the gastricglands are opened. Phytobezoars are formed in abnormalcondition and composed of indigestible food fibers such ascellulose which occur in fruits and vegetables including celery,pumpkin, prunes, raisins, leeks, beets, persimmons andsunflower-seed shells14. It is obvious from the study ofHafeez et al.15 that Hystrix indica is largely herbivorous in dietand serious pest of seasonal vegetable, fruit, trees, crops andgrasses. In crested porcupine (Hystrix cristata) structure ofphytobezoars have found in the stomach16. In agreement withthose studies, it is likely that phytobenzoars-like structure inthe surface of mucosa stomach of sunda porcupine wasformed likely in normal condition since, it found in all sampleof both sexes used in the present study. The contents ofphytobenzoars covered the mucosal surface of sundaporcupine stomach were confirmed by using the AB-PASand identified its contents were neutral mucopolysaccarideas indicated by their pink or magenta colors. Themucopolysaccharide represent as buffer system protectionagainst acidic substance and may be important for lubricationof mucosa17 from auto digestion18. The current study differwith in simple stomach19, in human stomach20 and in rodent21.

The narrow cardiac glands of sunda porcupine arecomposed by mucous secreting glands consist of seroussecreting cells. The appearances of cardiac glands are similarto that in feline stomach by Al-Tikriti et al.22 but disagreementwith the appearance of mucous-secreating cells as in simplestomach described by Samuelson23. There is not differencebetween the histological structures of the fundic glands insunda porcupine stomach from the histological structure ofthe fundic glands in the simple stomach of domestic

mammals described by Chivers and Hladik24. The compositionof fundic glands in sunda porcupine stomach confirmed withBacha and Bacha25 in monogastric animals and the parietalcells have the same histological characteristic in the stomachof monogastric animals and ruminant as reported byRobinson et al.26. The parietal cells are the predominant maintype cells of the fundic glands, similar histological descriptionby Cormack19 and Wheater et al.20 in simple stomach andSmith et al.1 in monogastric animals. The appearance ofpyloric glands of sunda porcupine is similar to the descriptionof Wheater et al.20. The pyloric glands of sunda porcupine havepredominantly mucous cells with a few parietal cells, thisfinding agrees with De Barros Moraes et al.27 and they definedthe pyloric glands region containing dominant mucous cellswith a little number of parietal and chief cells. This studydisagrees with Robinson et al.26 who showed that the pyloricglands consist of mucous-secreting cells only and abruptdisappearance of parietal and chief cells marks the boundarybetween pyloric and fundic glands. In the sunda porcupine,parietal and chief cells gradually decrease their number anddisappear in the end of the pyloric gland region. There is studyexamines the food habits of the Indian crested porcupine(Hystrix indica) but no information about the structure ofstomach porcupine at present, so this is the first basic data onthe histological stomach structure of the porcupine, especiallysunda porcupine (Hystrix javanica).

The AB-PAS stain has been used to highlight themucus-secreting cells of the gastric mucosa. This study notedthat the surface lining cells of entire mucosa of rabbit stomachin both sexes that extend into gastric pits gave a strongreactivity with PAS which appear as pink or magenta colorand they are considered as mucus producing cells. Themucous is positive for PAS because it is a neutralpolysaccharide and positive for AB if it is acid polysaccharide17.The fundic glands gave a positive reaction with AB due to themain cells of fundic gland are parietal cells which its functionin manufacture and secretion of hydrochloric acid and gastricproenzyme, respectively27, the finding of this study disagreeswith Leblond and Stevens28 in rat, Menzies29 in rabbits, Looand Wong30 in toad, Mouly and Rao31 in buffalo, Cormack19 inhuman and Sheahan and Jervis32 in laboratory animals. Ingastric and pyloric glands regions the gastric and pyloricglands gave a positive reaction with PAS since thepredominant cells are mucous secreting cells, some glands inpyloric region contain a very few No. of parietal cells. Thisresult is agree with Sheahan and Jervis32 in human Mori andSforzi16 in reptile. The difference in the reactivity of the gastricglands of rabbit with PAS may by a reflection of differentfunctions of these glands in stomach of sunda porcupine.

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CONCLUSION

In conclusion, the tunica mucosa stomach ofHystrix javanica consists of three lamina, lamina epithelialismucosa, lamina propria mucosa and lamina muscularismucosa. The mucosa stomach of sunda porcupine was dividedinto three gland regions, richly neutral mucopolysacaridesproducing cells of cardiac, acidic mucopolysacaridesproducing cells of fundic, mixed acid and neutralmucopolysacarides producing cells of pyloric glands regions.All of the mucosa stomach covered by neutralmucopolysacarides of thick phytobenzoars-like structures.

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