Type VII collagen is one of the major structural components of the corneal epithelial adhesion complex. Using the immunogold technique combined with indirect immunofluorescence analysis, the fine structural distribution of type VII collagen was studied in the corneas obtained from 5 enucleated hyman eyes (age range, 1-77 years) including one pathologic cornea from graft rejection. The findings on normal cornea corroborated the results from previous studies. In pathologic cornea from graft rejection, type VII collagen antibodies generated linear and irregular patchy fluorescence staining along the epithelial-stromal interface and immunogold binding to type VII collagen mainly occurred within the undulating lamina densa, more densealy distributed anchoring plaques and anchoring fibrils. The distribution of type VII collagen in pathologic human cornea from graft rejection is similar to normal human cornea. But, in pathologic cornea, type VII collagen is more densely distributed in superficial stroma and forms more extended anchoring network, which may be derived from the increased secretion of the type VII collagen due to the activated basal epithelial cell during healing process.
Antibodies ; Collagen Type VII* ; Cornea* ; Epithelial Cells ; Fluorescence ; Fluorescent Antibody Technique, Indirect ; Graft Rejection ; Humans ; Immunohistochemistry

OBJECTIVETo localize the attractin protein in the testis and epididymis of mature male rats.

METHODSTestes and epididymides were obtained from mature male Sprague dawley rats (n = 20). Tissues were fixed and prepared for immunohistochemical (IHC) and indirect immunofluorescent (IIF) assay, carried out with antiserum against rat attractin.

RESULTSIn the testis of the male rat, there was distinct immunopositive staining on cell membrane and cytoplasm within Leydig cells, primitive spermatogonia, primary spermatocytes, spermatids, Sertoli cells, and peritubular myoid cells. In the epididymis, including caput, corpus, cauda, there was no definitive immunopositive staining within the efferent ductule and epididymal duct.

CONCLUSIONAttractin is expressed in the male rat reproductive system and localized within Leydig cells and germ cells. It may be invoved in acting on the reproductive system. And its physiological function has yet to be further studied.

Animals ; Epididymis ; chemistry ; Fluorescent Antibody Technique, Indirect ; Immunohistochemistry ; Male ; Membrane Proteins ; analysis ; Rats ; Rats, Sprague-Dawley ; Testis ; chemistry

No abstract available.
Chlamydia* ; Fluorescent Antibody Technique, Indirect*

A 66-year-old man with a history of multiple transurethral resections for recurrent bladder tumors, staged as Ta according to the International Union Against Cancer staging guidelines, presented with a complaint of dry cough. A round nodule with a diameter of 7.5 cm was detected in the lung by chest computed tomography, and a video-assisted thoracoscopic lobectomy was performed. Pulmonary metastasis of recurrent bladder cancer was diagnosed by immunohistochemistry staining for the urothelium-specific protein uroplakin Ia. Subsequently, 2 cycles of systemic chemotherapy were administered. Two and a half years after treatment, no recurrence of pulmonary lesions has been detected. A combination of complete resection of pulmonary lesions and systemic chemotherapy may result in a good prognosis for patients with non-muscle-invasive bladder cancer.
Cough ; Fluorescent Antibody Technique, Indirect ; Humans ; Immunohistochemistry ; Lung ; Neoplasm Metastasis ; Neoplasm Staging ; Prognosis ; Recurrence ; Solitary Pulmonary Nodule ; Thorax ; Urinary Bladder ; Urinary Bladder Neoplasms ; Uroplakin Ia

BACKGROUND AND PURPOSE: Cutaneous nerve biopsies based on two-dimensional analysis have been regarded as a creditable assessment tool for diagnosing peripheral neuropathies. However, advancements in methodological imaging are required for the analysis of intact structures of peripheral nerve fibers. A tissue-clearing and labeling technique facilitates three-dimensional imaging of internal structures in unsectioned, whole biological tissues without excessive time or labor costs. We sought to establish whether a tissue-clearing and labeling technique could be used for the diagnostic evaluation of peripheral neuropathies. METHODS: Five healthy individuals and four patients with small-fiber neuropathy (SFN) and postherpetic neuralgia (PHN) were prospectively enrolled. The conventional methods of indirect immunofluorescence (IF) and bright-field immunohistochemistry (IHC) were adopted in addition to the tissue-clearing and labeling method called active clarity technique-pressure related efficient and stable transfer of macromolecules into organs (ACT-PRESTO) to quantify the intraepidermal nerve-fiber density (IENFD). RESULTS: The mean IENFD values obtained by IF, bright-field IHC, and ACT-PRESTO in the healthy control group were 6.54, 6.44, and 90.19 fibers/mm², respectively; the corresponding values in the patients with SFN were 1.99, 2.32, and 48.12 fibers/mm², respectively, and 3.06, 2.87, and 47.21 fibers/mm², respectively, in the patients with PHN. CONCLUSIONS: This study has shown that a tissue-clearing method provided not only rapid and highly reproducible three-dimensional images of cutaneous nerve fibers but also yielded reliable quantitative IENFD data. Quantification of the IENFD using a tissue-clearing and labeling technique is a promising way to improve conventional cutaneous nerve biopsies.
Biopsy ; Fluorescent Antibody Technique, Indirect ; Humans ; Imaging, Three-Dimensional ; Immunohistochemistry ; Methods ; Nerve Fibers ; Neuralgia, Postherpetic ; Peripheral Nerves ; Peripheral Nervous System Diseases ; Prospective Studies

This study was performed to investigate the morphology of the enteric nervous system and interstitial cells of Cajal (ICC) in the murine ileum. The PGP9.5-like immunoreactive (PGP9.5-LI) neurons and the c-Kit-like immunoreactive (c-Kit-LI) ICCs were stained by indirect immunofluorescence method and were observed under the confocal laser scanning microscopy. According to three dimensional reconstruction study, it was found that the PGP9.5-LI neurons and the c-Kit-LI ICCs were widely distributed in the intestinal wall : (1) In circular muscle layer, PGP9.5-LI nerve fibers were paralell to circular muscle layer. (2) In the myenteric plexus, the PGP9.5-LI nerve were closely apposed to the adjacent PGP9.5-LI nerve, constituting the networks. (3) In double-labeling immunohistochemistry using anti-PGP9.5 and anti-c-Kit antibodies, the c-Kit-LI networks encircled around the neural strands. The characteristic arrangement of the PGP9.5-LI enteric nervous system and the ICC containing c-Kit-positive cells provide a morphological basis upon the mechanism regulating gastro-intestinal motility and the pathogenesis of gatro-intestinal disorders.
Animals ; Antibodies ; Enteric Nervous System* ; Fluorescent Antibody Technique, Indirect ; Ileum* ; Immunohistochemistry ; Interstitial Cells of Cajal* ; Mice ; Microscopy, Confocal* ; Myenteric Plexus ; Nerve Fibers ; Neurons

OBJECTIVETo study the effect of activated microglia grafting on rats' hind limb motor function recovery after spinal cord injury.

METHODSMicroglia were separated from primary culture and subcultured for 3 generations. Lipopolysaccharide was added to the culture medium with the terminal concentration of 10 microl/L for microglia activation 3 days before transplantation. Totally 80 adult Wistar rats were divided into transplantation group and control group, with 40 rats in each group. Spinal cord injury model of rats was set by hitting onto the spinal cord using a modified Allen impactor. With a 5 microl micro-syringe, the activated microglia suspension was injected into the injured area 7 days after the first operation. Basso, Beattie and Bresnahan (BBB) scoring for hind limb motor function was taken on the 1st, 7th, 14th, 21st, and 28th day after microglia transplantation, and 8 rats were sacrificed at each time point mentioned above, respectively. Frozen sections of the spinal cord were made for haematoxylin-eosin (HE) and Naoumenko-Feigin stainings. SPSS 11.0 software was used for statistical analysis.

RESULTSBBB scores for hind limb motor function on the 14th, 21st, and 28th day were significantly higher compared with the control group. Most liquefaction necrosis areas disappeared and only a few multicystic cavities surrounded by aggregated microglia remained in the transplantation group. Naoumenko-Feigin staining for microglia showed that the transplantation group had significantly more positive cells (P < 0.05).

CONCLUSIONSGrafting of activated microglia into the injured spinal cord can significantly promote the hind limb motor function recovery in rats with spinal cord injury and reduce the size of liquefaction necrosis area. The extent of lower limb motor function improvement has a positive correlation with the number of aggregated microglia.

Animals ; Antigens, CD ; analysis ; Antigens, Differentiation, Myelomonocytic ; analysis ; Cells, Cultured ; Flow Cytometry ; Fluorescent Antibody Technique, Indirect ; Hindlimb ; physiopathology ; Immunohistochemistry ; Microglia ; transplantation ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Spinal Cord Injuries ; immunology ; physiopathology ; therapy

OBJECTIVETo examine the expression of heat shock protein 25 (HSP-25) in dental rat follicles in vivo and in vitro in order to investigate the possible effect of HSP-25 on cell proliferation and alkaline phosphatase (ALP) activity.

METHODSThe expression of HSP-25 in mandibles of postnatal rats from day 1, 3, 5, 7, 9, 11 was examined by immunohistochemistry in vitro, the expression of HSP-25 in the dental follicle cells was detected by the indirect immunofluorescence method. Methyl thiazolyl tetrazolium (MTT) assay, flowcytometry and ALP assay were used to detect the effect of HSP-25 on rat dental follicles.

RESULTSHSP-25 expression was absent or weak in rat dental follicle cells at early postnatal stage and present from day 5 till day 11. HSP-25 was detected in the cytoplasm of cultured dental follicle cells. MTT results showed no effect could be detected on dental follicle cell proliferation after stimulation of different concentrations of HSP-25 on day 1, 2, 3. Flowcytometry results also exhibited no difference in cell cycles after stimulation of HSP-25 at 0 microg/L and 100 microg/L. HSP-25 at a concentration of 50 microg/L and 100 microg/L could up-regulate the ALP activity on day 9.

CONCLUSIONSExpression of HSP-25 increases chronologically in the rat dental follicle cells. HSP-25 locates in the cytoplasm of cultured rat dental follicle cells. HSP-25 has no effect on the proliferation of dental follicle cells, however it can up-regulate the ALP activity.

Alkaline Phosphatase ; metabolism ; Animals ; Cell Proliferation ; Cells, Cultured ; Dental Sac ; metabolism ; Flow Cytometry ; Fluorescent Antibody Technique, Indirect ; HSP27 Heat-Shock Proteins ; physiology ; Immunohistochemistry ; Rats ; Tetrazolium Salts ; Thiazoles ; Up-Regulation

No abstract available.
Diagnosis* ; Fluorescent Antibody Technique, Indirect* ; Herpes Simplex* ; Simplexvirus*

The pathophysiology of glandular dysfunction in Sjögren's syndrome (SS) has not been fully elucidated. Previously, we reported the presence of autoantibodies to AQP-5 in patients with SS, which was associated with a low resting salivary flow. The purpose of this study was to investigate the presence of anti-AQP1 autoantibodies. To detect anti-AQP1 autoantibodies, cell-based indirect immunofluorescence assay was developed using MDCK cells that overexpressed human AQP1. By screening 112 SS and 52 control sera, anti-AQP1 autoantibodies were detected in 27.7% of the SS but in none of the control sera. Interestingly, the sera that were positive for anti-AQP1 autoantibodies also contained anti-AQP5 autoantibodies in the previous study. Different from anti-AQP5 autoantibodies, the presence of anti-AQP1 autoantibodies was not associated with the salivary flow rate. Although anti-AQP1 autoantibodies are not useful as a diagnostic marker, the presence of autoantibodies to AQP1 may be an obstacle to AQP1 gene therapy for SS.
Aquaporin 1 ; Autoantibodies* ; Fluorescent Antibody Technique ; Fluorescent Antibody Technique, Indirect ; Genetic Therapy ; Humans ; Madin Darby Canine Kidney Cells ; Mass Screening