Small bowel perforation secondary to enteric Salmonella paratyphi A infection.

A patient of Pakistani-origin was admitted to Bradford Royal Infirmary, UK, following a 3-week history of cough, headache and general malaise. He had recently spent 10 weeks in Pakistan and on return had been diagnosed in the community with Swine flu. He developed abdominal pain and diarrhoea in the week prior to admission, and presented to hospital with fever, tachycardia and raised inflammatory markers. He deteriorated rapidly, developing signs of peritonism and Salmonella paratyphi A was grown from blood cultures. CT demonstrated a small volume of free fluid within the abdomen and the patient underwent laparotomy. A small bowel perforation was resected and a side to side anastomosis fashioned. Treatment with intravenous antibiotics was completed and the patient was discharged 9 days postoperatively.

Regeneration of articular cartilage--evaluation of osteochondral defect repair in the rabbit using multiphasic implants.

OBJECTIVE: To investigate whether two different multiphasic implants could initiate and sustain repair of osteochondral defects in rabbits. The implants address the malleable properties of cartilage while also addressing the rigid characteristics of subchondral bone. DESIGN: The bone region of both devices consisted of D, D-L, L-polylactic acid invested with hyaluronan (HY). The cartilage region of the first device was a polyelectrolytic complex (PEC) hydrogel of HY and chitosan. In the second device the cartilage region consisted of type I collagen scaffold. Eighteen rabbits were implanted bilaterally with a device, or underwent defect creation with no implant. At 24 weeks, regenerated tissues were evaluated grossly, histologically and via immunostaining for type II collagen. RESULTS: PEC devices induced a significantly better repair than untreated shams. Collagen devices resulted in a quality of repair close to that of the PEC group, although its mean repair score (19.0+/-4.2) did not differ significantly from that of the PEC group (20.4+/-3.7) or the shams (16.5+/-6.3). The percentage of hyaline-appearing cartilage in the repair was highest with collagen implants, while the degree of bonding of repair to the host, structural integrity of the neocartilage, and reconstitution of the subchondral bone was greatest with PEC devices. Cartilage in both device-treated sites stained positive for type II collagen and GAG. CONCLUSIONS: Both implants are capable of maintaining hyaline-appearing tissue at 24 weeks. The physicochemical region between the cartilage and bone compartments makes these devices well suited for delivery of different growth factors or drugs in each compartment, or different doses of the same factor. It also renders these devices excellent vehicles for chondrocyte or stem cell transplantation.

Characterization of Tamm-Horsfall protein in a rat nephrolithiasis model.

PURPOSE: The role of Tamm-Horsfall protein in calcium oxalate stone formation is controversial. It is unclear whether Tamm-Horsfall protein has a role in crystallization. If it does, does it act as an inhibitor or promoter of crystallization? To elucidate the nature of its involvement we characterized Tamm-Horsfall protein in a rat model of calcium oxalate nephrolithiasis by in vivo and in vitro techniques. MATERIALS AND METHODS: Calcium oxalate nephrolithiasis was induced in male Sprague-Dawley rats. The amino acid and carbohydrate composition of Tamm-Horsfall protein from normal rats and those with nephrolithiasis was determined. The Tamm-Horsfall protein gene and protein expression in the kidneys were examined by in situ hybridization and immunohistochemistry. Furthermore, the interaction of Tamm-Horsfall protein and calcium oxalate crystals was assessed by an in vitro crystal aggregation assay. RESULTS: Tamm-Horsfall protein from rats with nephrolithiasis was biochemically similar to that from normal rats. Although Tamm-Horsfall protein was associated with crystal deposits in the renal papillae of rats with nephrolithiasis, Tamm-Horsfall protein messenger RNA expression in the kidneys remained unchanged. In each group Tamm-Horsfall protein inhibited calcium oxalate crystal aggregation by 47%, indicating no change in functional capabilities. CONCLUSIONS: The results of this study indicate that urinary excretion, and the biochemical nature and functional capabilities of Tamm-Horsfall protein remain unchanged during experimental calcium oxalate nephrolithiasis. Although staining for Tamm-Horsfall protein was evident in the papillae of rats with nephrolithiasis, the site of Tamm-Horsfall protein synthesis remained cells of the thick ascending limbs of the loop of Henle.

The cytoplasmic domain of human FcgammaRIa alters the functional properties of the FcgammaRI.gamma-chain receptor complex.

The gamma/zeta-chain family of proteins mediate cell activation for multiple immunoglobulin receptors. However, the recognition that these receptors may have distinct biologic functions suggests that additional signaling elements may contribute to functional diversity. We hypothesized that the cytoplasmic domain (CY) of the ligand binding alpha-chain alters the biological properties of the receptor complex. Using macrophage FcgammaRIa as a model system, we created stable transfectants expressing a full-length or a CY deletion mutant of human FcgammaRIa. Both receptors functionally associate with the endogenous murine gamma-chain. However, we have established that the CY of FcgammaRIa directly contributes to the functional properties of the receptor complex. Deletion of the FcgammaRIa CY leads to slower kinetics of receptor-specific phagocytosis and endocytosis as well as lower total phagocytosis despite identical levels of receptor expression. Deletion of the CY also converts the phenotype of calcium independent FcgammaRIa-specific phagocytosis to a calcium-dependent phenotype. Finally, deletion of the CY abrogates FcgammaRIa-specific secretion of interleukin-6 but does not affect production of interleukin-1beta. These results demonstrate a functional role for the CY of FcgammaRIa and provide a general model for understanding how multiple receptors that utilize the gamma-chain can generate diversity in function.

Reactive oxygen intermediates enhance Fc gamma receptor signaling and amplify phagocytic capacity.

Receptors for the Fc region of IgG (Fc gamma R) mediate internalization of opsonized particles by human neutrophils (PMN) and mononuclear phagocytes. Cross-linking of Fc gamma R leads to activation of protein tyrosine kinases and phosphorylation of immunoreceptor tyrosine-based activation motifs (ITAMs) within Fc gamma R subunits, both obligatory early signals for phagocytosis. Human PMN constitutively express two structurally distinct Fc gamma R, Fc gamma RIIa and Fc gamma RIIIb, and can be induced to express Fc gamma RI by IFN-gamma. We have previously shown that stimulation of PMN through Fc gamma RIIIb results in enhanced Fc gamma RIIa-mediated phagocytic activity that is inhibited by catalase. In the present study, we have tested the hypothesis that reactive oxygen intermediates (ROI) have the capacity to regulate Fc gamma R responses and defined a mechanism for this effect. We show that H2O2 augmented phagocytosis mediated by Fc gamma RIIa and Fc gamma RI in PMN and amplified receptor-triggered tyrosine phosphorylation of Fc gamma R-associated ITAMs and signaling elements. Generation of endogenous oxidants in PMN by cross-linking Fc gamma RIIIb similarly enhanced phosphorylation of Fc gamma RIIa and Syk, a tyrosine kinase required for phagocytic function, in a catalase-sensitive manner. Our results provide a mechanism for priming phagocytes for enhanced responses to receptor-driven effects. ROI generated in an inflammatory milieu may stimulate quiescent cells to rapidly increase the magnitude of their effector function. Indeed, human monocytes incubated in the presence of stimulated PMN showed oxidant-induced increases in Fc gamma RIIa-mediated phagocytosis. Definition of the role of oxidants as amplifiers of Fc gamma R signaling identifies a target for therapeutic intervention in immune complex-mediated tissue injury.

A prospective randomised controlled study of the use of ranitidine in patients with gastric cancer. Yorkshire GI Tumour Group.

BACKGROUND: Does the use of the histamine H2 receptor antagonist ranitidine improve the outcome of patients with gastric cancer? PATIENTS: A total of 222 patients with gastric cancer who had received radical or palliative resection or who were deemed inoperable at presentation. SETTING: Hospitals within Yorkshire, the participating clinicians being members of the Yorkshire GI Tumour Group. METHODS: A multicentre prospective randomised double blind trial comparing ranitidine 150 mg twice daily with placebo twice daily was undertaken. The principal outcome measures were survival and survival excluding those who died within 30 days of operation. RESULTS: The median survival (95% confidence intervals) was 331 (232 to 393) days for patients in the ranitidine group compared with 187 (143 to 269) for those in the placebo group. The difference in survival was not statistically significant (p = 0.225). When patients who died within 30 days of operation were excluded (21 in the placebo group, 15 in the ranitidine group), the difference in survival remained not significant (p = 0.358). No subgroup could be identified who significantly benefited from treatment, but for patients with stage VIa cancer the median survival was 134 days with placebo compared with 313 days with ranitidine (p = 0.073). CONCLUSION: This study does not show significant benefit from the use of ranitidine for gastric cancer but further larger studies may be indicated.

Biochemical and quantitative analysis of Tamm Horsfall protein in rats.

The involvement of Tamm Horsfall protein (THP) in nephrolithiasis is currently under investigation in several laboratories. Although rat is a commonly used species as an in vivo model for such studies, there is only limited information available about the biochemical properties and excretion profile of THP in normal rats. In order to characterize rat THP, we purified and analyzed normal male rat THP, and compared it with normal human male urinary THP by gel electrophoresis. Both THPs migrated at approximately 90 KDa, and stained similarly for protein (Coomassie blue) as well as carbohydrates (periodic acid Schiff reagent). Compositional analysis revealed that rat THP was largely similar to human THP in amino acid and carbohydrate contents but showed differences in the individual sugar components from other mammals. There was considerable variation in the day-to-day urinary excretion of THP in normal rats, with values ranging from 552.96 micrograms to 2865.60 micrograms and a mean value of 1679.54 micrograms per 24 h. It was concluded from this study that rat THP did not contain any unusual biochemical components and was primarily similar to human THP in composition and mean urinary concentration.

Immunocytochemical localization of Tamm-Horsfall protein in the kidneys of normal and nephrolithic rats.

Studies using in vitro systems have indicated that Tamm-Horsfall protein (THP) can interact with calcium oxalate (CaOx) crystals during kidney stone formation. However, information regarding the nature of its participation in this process remains controversial and unclear. In order to better understand the putative interaction of THP and crystals in vivo, we compared the localization of THP in normal rats and in chronic and semi-acute rat models of nephrolithiasis. In these rats, CaOx crystal deposits were induced in the kidneys by administering ethylene glycol (EG) in drinking water. The formation of CaOx mono- and dihydrate aggregates in the urine was confirmed by scanning electron microscopy. Immunohistochemical localization, as well as protein A-gold labeling at the ultrastructural level, demonstrated that in addition to its normal distribution, THP specifically associated with the renal crystal deposits. The THP-containing, organic matrix-like material consisted of a fine, fibrillar meshwork surrounding individual crystals and their aggregates. In addition, THP also appeared in the papilla, where it is normally absent, concurrent with the appearance of crystal deposits in the kidneys. These observations indicate that in nephrolithic rats the normal localization of THP is altered. Such an alteration may indicate an important physiological event related to crystal aggregation and kidney stone formation.

Localization of tamm-horsfall protein and osteopontin in a rat nephrolithiasis model.

The possibility of more than one urinary protein being simultaneously associated with calcium oxalate (CaOx) crystallization in vivo was investigated by examining the localization of Tamm-Horsfall protein (THP) and osteopontin (Opn) in a rat model of nephrolithiasis. CaOx crystal deposits were induced in male Sprague-Dawley rats by feeding 0.75% ethylene glycol in drinking water. THP and Opn were localized on kidney sections by immunoperoxidase technique, using specific polyclonal antibodies. When only occasional crystal deposits were seen in the kidney, THP showed a similar to normal pattern of distribution, with positive staining in the thick ascending limbs of the loop of Henle. Opn was localized in some nephrons in the thin limb of loop of Henle and on the papillary surface in the calyceal fornix. In contrast, in samples with a significantly increased number of deposits in the kidneys, the staining for both THP and Opn was strikingly enhanced and altered, with positive staining around the crystals as well as abnormal localization in the papilla. Interestingly, the occurrence of Opn was, however, more consistent than that of THP. This is a first study showing that in this nephrolithiasis model, normal localization of THP and Opn is altered and they are closely and concurrently associated with crystal deposits in vivo.

Characterization of a bombesin/gastrin-releasing peptide receptor on a human gastric-cancer cell line.

This study examined the expression of receptors of the bombesin (BBS) family in human gastric-cancer cell lines. Of 5 cell lines screened, only one, St42, demonstrated specific binding sites for 125I-Tyr4-BBS, which have been further characterized. This binding was saturable, and temperature- and time-dependent. Scatchard analysis of displacement data performed at 37 degrees C revealed 2 binding sites: a high-affinity, low-capacity site (KD = 0.13 nM, Bmax = 1500 sites/cell) and a lower-affinity, higher-capacity site (KD = 11 nM, Bmax = 35,000 sites/cell); the latter was lost when internalization of peptide was prevented, suggesting that it may be an artefact. Displacement assays with gastrin-releasing peptide (GRP) and neuromedin B (NMB) revealed that the receptor was of the GRP-preferring sub-type (GRP IC50 = 0.35 nM; NMB IC50 = 112 nM). Co-valent cross-linking of 125I-Tyr4-BBS to the receptor demonstrated the presence of a single band corresponding to a molecular weight of 37 to 44 kDa on SDS-PAGE, similar to that of the cloned GRP receptor protein core. G-protein linkage of this receptor was demonstrated by selective inhibition of 125I-Tyr4-BBS binding by guanosine nucleotides. The binding of BBS to the receptor resulted in a rise in intracellular calcium. Three of four structurally distinct BBS antagonists bound to the receptor with high affinity, but [DPhe12, Leu14]-bombesin did not cause any displacement of 125I-Tyr4-BBS even at 10 mM. The functional significance of GRP receptors on human gastric-cancer cells is as yet unknown, but further studies may determine whether such receptors have importance in the therapy of gastric cancer.

Structure of rat kidneys following microwave accelerated fixation.

In contrast to fixation of tissue in externally heated fixative, microwave-irradiation can generate uniform internal heat, which is of utmost importance for successful fixation of biological tissue. To evaluate the effectiveness of microwave-accelerated chemical fixation, we compared the structure of rat kidney fixed by a conventional method and a microwave-accelerated method, by scanning and transmission electron microscopy. Following perfusion, rat kidney pieces 1-2 mm in size were irradiated in Karnovsky's fixative in a domestic Amana microwave oven, till the temperature of the fixative reached 45-50 degrees C. For conventional fixation, tissue pieces were fixed overnight at room temperature in the same fixative. Both types of samples were processed further for electron microscopy using identical protocols. The microwave fixed samples showed excellent preservation of structure comparable to the samples fixed by the conventional method. Glomeruli and the renal tubules showed normal morphology with no cellular swelling. The cytoplasm and nuclear matrix of the epithelial cells was uniformly dense. Other fixation sensitive organelles like mitochondria and Golgi apparatus showed superior preservation with continuous membranes. These results demonstrate that microwave accelerated chemical fixation results in excellent preservation of tissue structure, reduces processing time significantly and is therefore a practical alternative to conventional protocols.