Specific Enzyme-linked Immunosorbent Assay System Research Articles

Influence of Abutments Surface (machined vs. laser-microgrooved) in Soft-tissue Response During One Year of Function: Clinical and Biochemical Outcomes of a RCT with Split-Mouth Design

The purpose of this study was to evaluate peri-implant soft tissue response by assessing IL-6, IL-1b and MMP-8 levels in peri-implant crevicular fluid (PICF) around machined vs. laser-microgrooved implants/healing/prosthetic abutments during 1 year of function. Twenty-four patients each received 2 one-stage implants in a split mouth design on the same jaw. In each patient, one implant, one immediate healing, and one prosthetic abutment with a machined surface (M group), and one implant, one immediate healing abutment and one prosthetic abutment with a laser-microgrooved surface (LMS group) were used. PICF sampling, pocket probing depths (PPD) and bleeding on probing (BOP) were assessed at 1, 3, and 12 months. IL-6, IL-1b and MMP-8 levels were determined by specific enzyme-linked immunosorbent assay systems (ELISA). Repeated measure ANOVA was used to run comparisons with groups and between groups months at 1, 3, and 12 months. At 3 and 12 months, the LMS group showed significantly lower PD, BOP and IL-6, IL-1β and MMP-8 levels than the M group (P<.05). This study suggests the presence of more remodeling and/or inflammatory phenomena around implants/abutments with a machined surface than around implants/abutments with a laser-microgrooved surface.

Establishment of a novel specific ELISA system for rat N‐ and C‐ERC/mesothelin. Rat ERC/mesothelin in the body fluids of mice bearing mesothelioma

Mesothelioma is a type of malignant tumor that most commonly arises from the pleural or peritoneal membrane and is usually associated with previous exposure to asbestos. In humans, ERC/mesothelin is expressed on the normal mesothelium and in some cancers such as mesothelioma or ovarian carcinoma. Recently, several enzyme-linked immunosorbent assay (ELISA) systems for ERC/mesothelin have been developed, the reported usefulness of which has been assessed and demonstrated as a diagnostic tool. However, the basic roles or physiological functions of, and relationship between, ERC/mesothelin and asbestos exposure-mediated carcinogenesis remain to be resolved. In order to elucidate the precise mechanism, animal models of mesothelioma are desperately needed. In this study, we consider the development of a novel specific ELISA system for not only rat N-ERC/mesothelin but also C-ERC/mesothelin, and the first data on the presence of rat ERC/mesothelin in the body fluids of rat malignant mesothelioma-bearing nude mice. The transplanted mice have revealed the higher concentrations of rat N-ERC/mesothelin in the blood and ascites than C-ERC/mesothelin. We hope these novel ELISA systems are useful in the rat model system to clarify the mechanism of asbestos-induced carcinogenesis and to develop new effective drugs for mesothelioma.

Ischemic cerebral tissue and MCP-1 enhance rat bone marrow stromal cell migration in interface culture.

Intravascular administration of bone marrow stromal cells (MSCs) restores function in animal models of neural injury and neurodegeneration. Adult MSCs administered intravenously to rat migrate and express neural phenotypes in ischemic brain. The aim of the present study was to investigate the mechanisms targeting MSC migration into the ischemic brain. Monocyte chemoattractant protein-1 (MCP-1), a chemoattractant factor, was measured in rat ischemic brain at various time points after middle cerebral artery occlusion (MCAo) using a specific enzyme-linked immunosorbent assay system (ELISA). In addition, using a microchemotaxis chamber, we measured whether ischemic brain tissue extracts induce migration of MSCs and whether brain tissue extracts incubated with antibodies against MCP-1 reduce MSC migration. Our data indicate that ischemic brain MCP-1 levels significantly increase from 6 hours, peak at 48 hours after MCAo (p < 0.05), and thereafter gradually decrease. Brain tissue extract at 6 hours, 24 hours, and 48 hours after MCAo significantly increase MSC migration across the membrane of the microchemotaxis chamber compared to normal tissue (p < 0.05). However, when the ischemic brain tissue extracts are incubated with antibody against MCP-1, MSC migration is significantly reduced at 24 hours and 48 hours after MCAo compared to extracts without this antibody (p < 0.05). Our data suggest that MCP-1 contributes to MSC migration into the ischemic brain tissue environment.

Characterization of microglial cells and their response to stimulation in an organotypic retinal culture system

An organotypic culture system of the early postnatal rat retina was developed to study microglial activation within a tissue environment. One day after tissue preparation, microglial cells of the ganglion cell/nerve fiber layer revealed features of activation. Cells acquired an ameboid morphology as revealed by Bandeiraea simplicifolia lectin staining. Proliferation-as revealed by Ki67 immunocytochemistry-resulted in higher cell densities. In the supernatant, tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and monocyte chemoattractant factor-1 (MCP-1) were detected by using specific enzyme-linked immunosorbent assay systems, activated microglia being the most likely source of their release. After 6 days in vitro (div), microglial cells regained their resting morphology, and cell counts returned to control levels. Concomitantly, the release activity decreased to undetectable levels. When slices were treated at this later stage of cultivation (>6 div) with bacterial lipopolysaccharide (LPS; 100 ng/ml for 24 hours), microglial cells became activated, as revealed by a change in morphology. In parallel, the LPS treatment also resulted in high levels of TNF-alpha, IL-6, and MCP-1 in the culture medium. Both the release from the tissue and the morphological changes of the microglia were reversible. Seventy-two hours after LPS removal, only microglia with ramified morphology were found, and release activities returned to baseline. These data suggest that the organotypic culture of the retina is a useful model for studying microglial activation from its resting form.

Crossline levels in serum and erythrocyte membrane proteins from patients with diabetic nephropathy

Crossline is one of the structurally defined adducts of advanced glycation endproducts (AGEs) which has both a crosslink and fluorescence similar to AGE-protein in vivo. Crossline was measured in serum and erythrocyte membrane proteins (EMP) from 52 type 2 diabetic patients using a specific enzyme-linked immunosorbent assay system. Serum and EMP crossline levels in the diabetic patients were significantly higher than those in normal control. The patients with advanced diabetic nephropathy (serum creatinine levels of more than 1.2 mg/dl) had markedly elevated serum crossline levels compared to those with moderate diabetic nephropathy (clinical proteinuria) (180.7±51.7 vs. 71.8±18.4 pmol/ml; P<0.01). On the other hand, there were no significant differences in EMP crossline levels between the two. EMP crossline levels in the patients with moderate diabetic nephropathy (8.8±2.9 pmol/mg protein) and those with advanced diabetic nephropathy (9.7±3.0 pmol/mg protein) were significantly higher than those without clinical proteinuria (6.4±1.9 pmol/mg protein; P<0.01). The present study demonstrated that EMP crossline levels were associated with the presence of nephropathy in patients with type 2 diabetes mellitus. Serum crossline levels were significantly influenced by remaining renal function. The measurement of crossline from a blood sample could provide us with important information for the study of clinical evaluation and pathogenesis of diabetic complications.

Differential production of MCP-1 and cytokine-induced neutrophil chemoattractant in the ischemic brain after transient focal ischemia in rats.

Chemokines have been shown to play an important role in leukocyte infiltration into ischemic lesions. Recently, the increased expression of monocyte chemoattractant protein-1 (MCP-1) and cytokine-induced neutrophil chemoattractant (CINC) was observed in experimental stroke models where infiltrated leukocytes were supposed to induce tissue injury, however, the protein level and time course of these chemokines have not been fully elucidated. Therefore, we analyzed the time-dependent production of MCP-1 and CINC in the rat brain after transient middle cerebral artery occlusion (MCAO) by means of specific enzyme-linked immunosorbent assay systems. The MCP-1 levels in the ipsilateral hemispheres increased from 6 h, peaked at 2 days, and thereafter gradually decreased. The peak MCP-1 concentration was 89.2+/-28.2 ng/g tissue wet weight (mean +/- SEM, n = 5, 49.3-fold greater than the contralateral value at the same time, P < 0.05), which is supposed to be high enough to exert its biological effects. In contrast, the maximum CINC concentration that corresponded to 2.9+/-0.7 ng/g tissue wet weight (mean +/- SEM, n = 5, 55.0-fold greater than the contralateral value at the same time, P < 0.05), was observed at 6 h. In addition, we confirmed the temporal profile of leukocyte subtypes that infiltrated into the ischemic brain, thus, neutrophil infiltration occurred at early stages (1-3 days), followed by massive infiltration of macrophages at later stages (2-7 days). These studies suggest that MCP-1 in cerebral ischemia actually plays a significant role in the migration of macrophages into the lesion and that the differential temporal production of these chemokines contributes to the regulation of infiltrated leukocyte subtypes.