الفهرس 
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Abstract
Small-bowel ulcers are commonly caused by Crohn’s disease, abdominopelvic radiotherapy, chemotherapy for cancer, mycotoxins, alcohol intake, tuberculous enteritis, or drug induction. The most common causes of drug-induced intestinal mucosal injury are NSAIDs that may cause ulcers and erosions predominantly in the jejunum and ileum. Cell-based therapies are widely used in tissue regeneration as well as treatment of various diseases. PBMC therapy plays an important role in tissue healing and repair. Furthermore, priming PBMCs with APS has been known to augment their efficacy via induction of angiogenesis and tissue regeneration. Also, misoprostol which is a PG analogue has been reported to be effective for the treatment of small intestinal mucosal ulcers and erosions. Aim of the work: This work was done to study the effect of PBMCs as well as APSprimed PBMCs on the induced ulcers in the ileal mucosa of adult male albino rats compared to misoprostol. Materials and Methods: This study was carried out on 70 adult male albino rats; 15 of them were used as donors and the other 55 were used as recipients. The recipient rats were divided into two main groups as follows: 1- Control group (group I): included 15 rats that were subdivided into three subgroups (five rats each): Subgroup I-A: kept without any interference. Then the animals were sacrificed after 2 weeks. Chapter VII Summary and conclusion 202 Subgroup I-B: received 0.5 ml phosphate buffered saline (PBS) (the vehicle used to suspend the primed mobilized PBMCs) once intravenously via tail vein. Then the animals were sacrificed after 2 weeks. Subgroup I-C: received 1 ml Tween-80 solution (the vehicle used to prepare misoprostol) orally. Then the animals were sacrificed after daily administration of the vehicle for 2 weeks. 2- Experimental group (group II): included 40 rats that were subdivided into four subgroups (10 rats each): Subgroup II-A: received a single oral dose of 25 mg/kg indomethacin dissolved in 1 ml Tween‐ 80 and 9 ml 0.9% NaCl after overnight fasting. The animals were sacrificed 6 hours after induction of ulcer. Subgroup II-B: received 100μg/kg Misoprostol (dissolved in 0.75% Tween-80 solution) orally once daily for 2 weeks, 24 hours after induction of ulcer. Subgroup II-C: injected with mobilized PBMCs (1×107) suspended in 0.5 ml PBS once intravenously via tail vein 24 hours after induction of ulcer. Then the animals were sacrificed 2 weeks after the injection. Subgroup II-D: injected with APS-primed mobilized PBMCs (1×107) suspended in 0.5 ml PBS once intravenously via tail vein 24 hours after induction of ulcer. The animals were sacrificed 2 weeks after the injection. In the current study, mononuclear cells were isolated from peripheral blood of donor rats after their subcutaneous injection with Filgrastim (Neupogen) at a dose of 100μg/kg once daily for three consecutive days. At the end of the third day, peripheral blood was collected via cardiac puncture then the PBMCs were isolated by density gradient centrifugation method. Additionally, PRP was separated from blood of donor rats by centrifugation then it was activated with calcium gluconate. After that, the activated-platelet content was centrifuged, and the supernatant was used for priming the mononuclear cells. At the end of the experiment, the animals were anesthetized by intraperitoneal injection of sodium pentobarbital at dose of 50 mg/kg and the ileum was carefully dissected and processed for histological and immunohistochemical examination. For light microscopic study, paraffin sections were stained with H&E, combined PAS-AB stain, and immunohistochemical staining was used for detection of PCNA. On the other hand, uranyl acetate and lead citrate were used to stain ultrathin sections for electron microscopic examination. Moreover, Image J software (National Institute of Health, Bethesda, Maryland, USA) was used for image analysis of the mean number of goblet cells in both villi and crypts in combined PAS-AB stained sections. The goblet cells were counted under a magnification power of (×200) in both villi and crypts. In each specimen, a total of 10 randomly microscopic fields were examined. Also, the number of PCNA positive nuclei was counted at a magnification power of (×400) in the crypt region (10 randomly microscopic fields were examined for each specimen per animal) and evaluated by using Image J software. Results: 1- Control group (group I): Examination of this group by light and electron microscopes revealed normal histological features of the ileum. 2- Experimental group (group II): *Subgroup II-A: H&E-stained sections of this subgroup revealed marked histological structural damage in the form of disturbance of normal villi and crypts architecture. The mucosa appeared with focal damage, atrophy of villi, damaged tips of villi with desquamation and loss of surface epithelial cells. Some villi appeared broad with subepithelial space while others appeared fused. Also, damaged and widening of crypts together with dispersed areas in the lamina propria, heavy cellular infiltration, congestion of blood vessels and discontinuity of the muscularis mucosa were also observed. Moreover, the cells lining the villi and crypts appeared separated with vacuolated and fragmented cytoplasm, fragmented nuclei, and darkly stained nuclei with perinuclear halo. Combined (PAS-AB)-stained sections of subgroup II-A showed apparently few goblet cells over the villi and crypts exhibiting PAS-AB positive reaction. Some goblet cells appeared completely depleted of mucous and are PAS-AB negative. Lost brush border of most enterocytes was also observed. Statistical results of the data collected by Image J analysis displayed highly significant decrease in the mean number of goblet cells when compared with control group. Additionally, PCNA immunostained sections of this subgroup showed weak positive reaction in the nuclei of many cells in the crypts. Statistical results of the data collected by Image J analysis revealed highly significant decrease in the mean number of PCNA positive nuclei when compared with control group. The EM examination confirmed the light microscopic results. Enterocytes appeared with focal loss of microvilli together with areas of short sparse microvilli and others with destroyed luminal surface. The goblet cells showed an apparent decrease in their secretory granules. Some granules appeared irregular in shape and others fused together with depletion of secretion. Also, Paneth cells showed disrupted microvilli, depletion of their secretory granules and others showed abnormal granules with feathery edges and fusion with each other. Furthermore, the cells revealed swollen mitochondria with destroyed cristae, dilated RER, cytoplasmic vacuolation and rarefaction, shrunken nuclei with condensed chromatin, corrugation of the nuclear membrane, and dilated perinuclear space. These were accompanied with inflammatory cellular infiltration and congested blood vessels in the lamina propria. *Subgroup II-B: H&E-stained sections of this subgroup showed focal improvement of villi and crypts architecture when compared to subgroup II-A. However, some villi still showed abnormal shape and others are short with irregular outline. Also, distortion and destruction of the epithelial cell lining of villi was observed. Some cells lining the villi and crypts still appeared with fragmented and vacuolated cytoplasm, as well as dilated intercellular spaces together with darkly stained nuclei and perinuclear halo in addition to congested blood vessels in the lamina propria. Combined (PAS-AB)-stained sections of subgroup II-B showed an apparent increase in the number of goblet cells exhibiting PAS-AB positive reaction when compared to subgroup II-A. Also, PAS positive reaction in the brush border of most enterocytes was observed. Statistical results revealed highly significant increase in the mean number of goblet cells when compared with subgroup (II-A). Additionally, there was strong positive immunoreaction for PCNA in the nuclei of many cells in the crypt region when compared to subgroup (IIA). Statistical results revealed highly significant increase in the mean number of PCNA positive nuclei when compared with subgroup (II-A) and significant increase in the mean number of PCNA positive nuclei when compared to control group. Examination of ultrathin sections of this subgroup revealed an apparent partial improvement as compared to subgroup II-A. However, some cells still showed dilated RER, destructed mitochondria, electron dense bodies, and cytoplasmic vacuoles. Some nuclei appeared fragmented, and others were shrunken with condensed chromatin, corrugation and indentation of nuclear membrane, and dilated perinuclear space in addition to dilated intercellular space. Some goblet cells still appeared with fusion of mucin secretory granules and depletion of secretion. Also, cellular infiltration and congested blood vessels in the lamina propria were observed. *Subgroup II-C: H&E-stained sections of this subgroup showed an apparent moderate improvement of villi and crypts architecture when compared to subgroup IIA. However, some villi appeared short and others with subepithelial space. Some cells lining the villi and crypts still appeared destructed with fragmented and vacuolated cytoplasm and others appeared with darkly stained nuclei. Cellular infiltration of the lamina propria was also observed. Combined (PAS-AB)-stained sections of subgroup II-C showed an apparent increase in the number of goblet cells exhibiting PAS-AB positive reaction when compared to subgroup II-A. Also, PAS positive reaction in the brush border of most enterocytes was observed. Statistical results revealed a highly significant increase in the mean number of goblet cells when compared with subgroup (II-A) and no significant difference in the mean number of goblet cells when compared with subgroup (II-B). Moreover, PCNA immunostained sections of this subgroup showed a strong positive reaction for PCNA in the nuclei of many cells in the crypt region when compared to subgroup (II-A) or control group. Statistical results revealed highly significant increase in the mean number of PCNA positive nuclei when compared with subgroup (II-A), significant increase in the mean number of PCNA positive nuclei when compared to control group and no significant difference in the mean number of PCNA positive nuclei when compared with subgroup (II-B). Examination of ultrathin sections of this subgroup revealed an apparent moderate improvement in most of the cells as compared to subgroup II-A. However, few cells showed cytoplasmic vacuolation and rarefaction, swollen destroyed mitochondria, dilated RER, and electron dense bodies as well as nuclei with irregular outline. *Subgroup II-D: H&E-stained sections of this subgroup showed the best improvement of ileal mucosal architecture as compared to other subgroups. Most of the villi and crypts showed nearly normal architecture with a nearly normal enterocytes, goblet cells and Paneth cells. However, few enterocytes still appeared with vacuolated cytoplasm. Combined (PAS-AB)-stained sections of subgroup II-D showed numerous goblet cells distended with mucous with an apparent strong PAS AB positive reaction. Strong PAS positive reaction in the well-defined brush border was also observed. Statistical results revealed a highly significant increase in the mean number of goblet cells when compared with subgroups (II-A, II-B& II-C) and no significant difference in the mean number of goblet cells when compared with control group. Furthermore, PCNA immunostained sections of this subgroup showed a strong positive reaction for PCNA in the nuclei of numerous cells in the crypt region when compared to other subgroups. Statistical results revealed the highest significant increase in the mean number of PCNA positive nuclei when compared with subgroups (II-A, II-B& II-C) as well as control group. Examination of ultrathin sections of this subgroup revealed an apparent marked improvement with nearly normal cells as compared to other subgroups. Enterocytes appeared with preserved regularly arranged microvilli, intact lateral interdigitations and junctional complexes and rounded to oval nuclei besides apparently normal elongated mitochondria and RER. Intraepithelial lymphocytes appeared with clear cytoplasm and abundant mitochondria. Additionally, apparently normal goblet cells were observed in between enterocytes with apical part distended with mucin granules and basal part containing compressed nucleus and RER. As regards Paneth cells, they appeared with basal nuclei and apical membrane bound electron dense homogenous secretory granules.