Strini Naidoo

Kinin receptors in human vascular tissue: their role in atheromatous disease.

Using samples of many human blood vessels, obtained at autopsy and specific antibodies directed to peptide sequences of the kinin B1 and B2 receptors, we demonstrate the localisation of these receptors within the human vascular system using standard immunolabelling techniques. In large elastic arteries and veins, kinin receptors are present only in the endothelial cells whereas in all muscular arteries and arterioles, these receptors are present in both the endothelial and smooth muscle cells. The identification of kinin receptors in human blood vessels confirms that kinins may modulate both vascular permeability and contractility. The incidental finding at histology, of patchy atheromatous disease in the coronary, femoral, vertebral and pericallosal arteries, assisted in elucidating the role of these receptors in the commonest disease affecting human blood vessels. Intense labelling for B1 receptors was observed in the endothelial cells, foamy macrophages, inflammatory cells and fibroblasts within the thickened intima of the plaque as well as in smooth muscle cells of the underlying tunica media. Immunoreactive B2 receptors were also observed in these cells but with reduced intensity. The intense immunolabelling of B1 receptors in these regions suggest that these may be induced by atheromatous disease and may have therapeutic importance for the B1 receptor antagonists.

Tissue kallikrein and kinins in renal disease

The renal kallikrein-kinin system is involved in sodium and water homeostasis, blood pressure regulation and inflammation. Tissue kallikrein and kinin levels were measured in the urine of patients with renal disease and in the urine of living related kidney donors prior to uninephrectomy who served as controls. Tissue kallikrein and kinin B1 and B2 receptors were immunolocalised by confocal microscopy in renal biopsy material from patients with renal disease and controls (fresh autopsy material and normal kidney tissue from nephrectomies for malignancy). Urinary tissue kallikrein excretion was significantly decreased in patients with mild renal disease (16.6 +/- 6.7 ng tissue kallikrein (TK)/ng protein; p < 0.05) and more markedly so (1.8 +/- 0.7 ng TK/microg protein; p < 0.01) in patients with severe renal failure requiring dialysis compared to normal controls (78.9 +/- 31.7 ng TK/microg protein). Basal kinin values were unchanged in patients with renal disease (14 +/- 0.8 ng/ml) compared to controls (13.3 +/- 0.56 ng/ml). In control kidney tissue kallikrein was immunolocalised in the distal connecting tubules and collecting ducts whereas decreased immunolabelling was observed with renal disease. Kinin B2 receptor labelling was present in the entire nephron in the normal control kidney but was reduced with renal disease. While kinin B1 receptor immunolabelling was not observed in the control kidneys, labelling of distal tubules and collecting ducts was noted in renal disease, suggesting an upregulation of B1 receptors in renal parenchymal disease.

Immunolocalization of bradykinin receptors on human synovial tissue

Kinins have been implicated in the pathogenesis of experimental and clinical inflammatory arthritis. Previous studies have reported increased amounts of plasma and tissue kallikreins in synovial fluid, raised kinin levels and an upregulation of kinin B2 receptors on synovial fluid neutrophils in rheumatoid arthritis. Bradykinin binding sites have been identified on human synovial cells by autoradiographic localization and Scatchard analysis. This study was undertaken to localize immunohistochemically kinin B1 and B2 receptors on human synovial tissue. Synovial tissue was obtained at the time of joint replacement surgery or arthroscopic synovectomy in six patients (two RA, two OA and two with avascular necrosis). Tissue sections were immunolabelled for kinin B1 and B2 receptors and viewed by light and confocal microscopy. No immunolabelling of the kinin receptors was observed in the method controls. In all patients labelling for kinin B2 receptors was observed in the synovial lining cells, fibroblasts and endothelial lining cells of blood vessels. There was no immunolabelling for kinin B1 receptors in all samples. These findings further support a role for the B2 receptors in joint diseases. There did not appear to be an induction of the kinin B1 receptor in human synovial tissue obtained from patients with chronic arthritis. However, further studies are required to assess the role of B1 receptors in active joint inflammation.

Kinin receptor status in normal and inflammed gastric mucosa.

No documented studies have been reported on the presence of B1 and B2 kinin receptors in the mammalian gastric mucosa. This first study aimed to immunolocalise sites of B1 and B2 kinin receptors in the human pyloric gastric mucosa and to evaluate its role in gastritis. Biopsies were obtained from patients with dyspepsia during endoscopic examination of the patient. The diagnosis and grading of the gastritis was performed on histological examination. Sections were immunostained for both B1 and B2 receptors using rabbit anti-human B1 and B2 kinin receptor antibodies. Control tissue was obtained from partial gastrectomy specimens, following surgical excision of the antrum for duodenal ulcers. The control antrum tissue showed strong immunoreactivity for kinin B2 receptors with positivity noted along the luminal border, at the base of the mucous and stem cells. The B1 receptor was not immunolocalised. Biopsies of all five patients with gastritis showed a decrease in immunolabelling of the B2 receptor and an induction of the B1 receptor especially in regenerating cells. In gastritis there is destruction of the normal mucosal glandular architecture with subsequent regeneration of the epithelial cells. The pyloric glands are infiltrated by acute inflammatory cells that cause crypt abscesses with loss of the epithelial cell membranes. This may explain the reduction in the immunolocalisation of the B2 kinin receptors and the induction of the B1 receptors in active gastritis. Follow up studies after treatment of the inflammation with a combination of B1/B2 kinin receptor antagonists are indicated.

The evaluation of tissue kallikrein in Helicobacter pylori-associated gastric ulcer disease

Helicobacter pylori (Hp) associated ulcer disease is a common form of gastric disorder involving mucosal damage and invasion of the mucosa by polymorphic inflammatory cells with concomitant changes in the epithelial cell structure. The bacteria are thought to adhere by specific junction zones to the epithelial cell surface resulting in the degeneration of the mucosal layer. Our study was undertaken to examine the relative status of tissue kallikrein (TK) in antral and fundic biopsies, endoscopically obtained from 10 patients suspected of having gastric disorders. For histological evidence of inflammation the tissue was stained with hematoxylin and eosin and classified as mild, active, chronic and chronic active gastritis. Hp infection was determined by Giemsa staining. For localisation of TK, slide-mounted tissue sections were subjected to PAP and immunofluorescent staining using a goat anti-human TK IgG antibody. The results revealed that in the antral control tissue, removed during partial antractomy, TK was immunovisualised along the luminal border of the deep pyloric glands. The surface epithelia and superficial glands showed no labelling. The fundic control tissue revealed an absence of TK in the superficial and surface epithelial glands, but was positive in the parietal cells. The fundic biopsy specimens showed similar immunoreactivity in these areas. By contrast, in the inflammed pyloric mucosa, there was a shift of TK localisation to the basal part of the glandular cells and there was also expression of TK in the superficial glands that showed histological evidence of regeneration. In the fundic biopsies there was no change observed in the sites of TK localisation (similar to control tissue). It was observed, that even though 8 of the 10 subjects exhibited Hp infection, the inflamed mucosa showed no discernable difference in the staining patterns between the infected and non-infected tissue sections. Our findings suggest an important role for a B1/B2 kinin antagonist in patients with gastritis.

Tissue kallikrein in transplant kidney

Literature survey, thus far, has shown a decrease in the excretion of urinary tissue kallikrein (TK) in transplant patients with a further reduction of the enzyme during episodes of acute rejection. The study aims were to compare, at cellular and subcellular levels, the localisation of tissue kallikrein in biopsies of the transplant kidney to autopsy derived normal renal tissue. Renal biopsies from eighteen transplant patients with deteriorating renal function were obtained. Immunolabelling for tissue kallikrein, using a polyclonal goat anti-TK, antibody raised against recombinant TK, was performed following routine enzymatic, immunofluorescence and electron microscopic techniques. In normal kidney tissue, TK was immunolocalised in the distal connecting tubules and collecting ducts. By comparison the renal transplant tissue showed a reduction in the intensity of label, but maintained the sites of localisation. In the sections examined by electron microscopy, although TK was confined mainly at the luminal side of the cell, some label was noted along the basolateral membranes. In the transplant kidneys, there was a reduction in the overall number of gold particles counted, which correlated with the decreased intensity observed on immunocytochemistry. In addition, there was a shift to a basolateral orientation of the immunolabel. Acute rejection is characterised by oedema, tubulitis and vasculitis. Destruction of the tubule cells and leakage of TK into the interstitial tissue space and the resultant effect of the formed kinins on renal capillary vasculature could explain the observed renal parenchymal oedema and transplant rejection.