Phenytoin desensitization monitored by antigen specific T cell response using carboxyfluorescein succinimidyl ester dilution assay

We evaluated drug-specific T cell responses in a patient with refractory partial seizures and paroxysmal kinesigenic choreoathetosis successfully treated with clinical desensitization to phenytoin. Drug-induced lymphocyte transformation test before desensitization was negative with a stimulation index of 130%. The frequencies and cytokine-producing phenotypes of phenytoin-specific T cells were examined simultaneously by using a carboxyfluorescein succinimidyl ester (CFSE) dilution assay. Before desensitization, the proportion of CFSElow CD4+ cells in whole CD4+ was 3.09%; 13.6% of CFSElow CD4+ cells were stained with anti-interferon gamma antibody. After desensitization, phenytoin-specific CFSElow CD4+ cells decreased to background level. These results indicate that CFSE dilution assay will be useful for the diagnosis and monitoring of drug hypersensitivity.     

Regeneration of defects in articular cartilage in rat knee joints by CCN2 (connective tissue growth factor).

CTGF/CCN2, a hypertrophic chondrocyte-specific gene product, possessed the ability to repair damaged articular cartilage in two animal models, which were experimental osteoarthritis and full-thickness defects of articular cartilage. These findings suggest that CTGF/CCN2 may be useful in regeneration of articular cartilage. INTRODUCTION: Connective tissue growth factor (CTGF)/CCN2 is a unique growth factor that stimulates the proliferation and differentiation, but not hypertrophy, of articular chondrocytes in vitro. The objective of this study was to investigate the therapeutic use of CTGF/CCN2. MATERIALS AND METHODS: The effects of recombinant CTGF/CCN2 (rCTGF/CCN2) on repair of damaged cartilage were evaluated by using both the monoiodoacetic acid (MIA)-induced experimental rat osteoarthritis (OA) model and full-thickness defects of rat articular cartilage in vivo. RESULTS: In the MIA-induced OA model, quantitative real-time RT-PCR assays showed a significant increase in the level of CTGF/CCN2 mRNA, and immunohistochemical analysis and in situ hybridization revealed that the clustered chondrocytes, in which clustering indicates an attempt to repair the damaged cartilage, produced CTGF/CCN2. Therefore, CTGF/CCN2 was suspected to play critical roles in cartilage repair. In fact, a single injection of rCTGF/CCN2 incorporated in gelatin hydrogel (rCTGF/CCN2-hydrogel) into the joint cavity of MIA-induced OA model rats repaired their articular cartilage to the extent that it became histologically similar to normal articular cartilage. Next, to examine the effect of rCTGF/CCN2 on the repair of articular cartilage, we created defects (2 mm in diameter) on the surface of articular cartilage in situ and implanted rCTGF/CCN2-hydrogel or PBS-hydrogel therein with collagen sponge. In the group implanted with rCTGF/CCN2-hydrogel collagen, new cartilage filled the defect 4 weeks postoperatively. In contrast, only soft tissue repair occurred when the PBS-hydrogel collagen was implanted. Consistent with these in vivo effects, rCTGF/CCN2 enhanced type II collagen and aggrecan mRNA expression in mouse bone marrow-derived stromal cells and induced chondrogenesis in vitro. CONCLUSION: These findings suggest the utility of CTGF/CCN2 in the regeneration of articular cartilage.     

Regeneration of endothelial cells after balloon denudation of the rabbit carotid artery and changes in responsiveness

The present experiments were carried out to determine the regrowth of endothelial cells (EC) after balloon denudation of the rabbit carotid artery and the changes in responsiveness of the artery with regenerated EC. Scanning electron microscopic findings revealed that 28.8% of the luminal surface was covered with regenerated EC at week 1. The regrowth of EC proceeded progressively, and a full lining was achieved at week 6. Regenerated EC were morphologically different from native ones; they were elongated (weeks 1 and 2) and irregularly oriented (weeks 4 and 6), and their numbers had significantly increased. Light microscopy revealed the intimal thickening and proliferation of smooth muscle cells. No accumulation of lipids in the vascular wall could be detected at any observation time. The experiments in an organ bath demonstrated that the altered appearance of EC was accompanied by depressed endothelium-dependent relaxations to acetylcholine, ADP and A23187. However, sodium nitroprusside-induced relaxation and contractile responses to noradrenaline, serotonin and histamine remained unchanged in the normal and denuded preparations, indicating that the dysfunction of the endothelium occurs at a time when the ability of the underlying vascular smooth muscle to relax or contract was unchanged. In addition, it is suggested that the impairment of the endothelium-dependent relaxation may be partly due to impairment of the synthesis and/or release of endothelium-derived relaxing factor(s) in EC.     

81mKr scintigraphic evaluation of hemodynamics in gynecologic malignancies under condition of angiotensin II-induced hypertension]

Transcatheter arterial infusion chemotherapy is one of the most useful therapeutic procedures for gynecologic malignancies. Recently, several reports have been published about Angiotensin II-induced hypertension chemotherapy and the efficacy of the method, but there have been no reports to evaluate an application for gynecologic malignancies. We evaluate the usefulness of the method for gynecologic malignancies demonstrating the changes of hemodynamics of the tumor using 81mKr scintigraphy. Thirteen patients with pathologically confirmed gynecologic malignancies were evaluated by angiography and continuous infusion of 81mKr via the catheter with and without Angiotensin II. At first, continuous infusion of 81mKr was performed under the superselective catheterization of the uterine artery. The radioactivities in the ROI were counted. Then, withdrew the catheter from the uterine artery to the internal iliac artery, and again continuously infused 81mKr and counted the radioactivities in the same ROI. Finally, keeping the catheter in the internal iliac artery, Angiotensin II and 81mKr were infused simultaneously. And counted the radioactivities. The radioactivities were highest when the catheter tip was placed in uterine arteries and lowest when the catheter tip was placed in internal iliac arteries. But radioactivities in the ROIs were definitely increased when Angiotensin II was used, even if the catheter tip was keeping in the internal iliac arteries. The optimal catheter position of transcatheter arterial chemotherapy for gynecologic malignancies is at proximal uterine artery. Since Angiotensin II-induced hypertension may increase blood flow of tumors, it seems to have indication for post-operative cases, highly advanced cases and cases with difficulties to perform superselective catheterization.(ABSTRACT TRUNCATED AT 250 WORDS)     

In vivo selection of tumorigenic subline from non-tumorigenic human gastric carcinoma cells: in relation to proliferative properties in vivo and in nude mice

Human gastric carcinoma cells from one of three long-term cultured cell lines (HPE-GAC-T) were injected into peritoneal cavities of BALB/c mice. The surviving celss in vivo were collected 3 days later. Following brief cultivation in vitro, those cells were reinjected into mice by the same route. This procedure was repeated 3 times. The cultured cancer cells recovered from the mice on the 3rd passage, at a 92.5% recovery rate, showed xenotransplantability in BALB/C nu/nu mice by subcutaneous injection. This subline (GAC-T.M-2) can be maintained in vitro but not in vivo while maintaining heterotransplantability. Three original cancer cell lines did not show tumorigenicity in nude mice. Animal passages by the same protocol failed to select tumorigenic sublines from the other cell lines (HPE-GAC-2 and -3). Factors affecting tumorigenic capacity of cancer cells in nude mice were studied in vivo and in vitro by comparing the properties of GAC-T.M-2 and parental cancer cells (GAC-T.O). Treatment of the hosts by injection of anti-asialoGM1 antibody or cyclophosphamide, adult thymectomy of BALB/c mice, and 400 rads whole body irradiation did not enhance the growth of either GAC-T.M-2 or -T.O cells. There was no detectable difference between in vitro growth properties of the original and variant cells at a rather high cell density. However, at a low cell density GAC-T.M-2 cells showed a higher cell growth rate and increased [3H] thymidine incorporation and possessed higher colony forming activity in the liquid medium than their parental cells. High dense expression of epidermal growth factor (EGF) receptors was evident equally in both GAC-T cells, however, GAC-T.M-2 cells were more sensitive to down-regulation by EGF in culture. Tumor cells of HPE-GAC-2 and -3 lines expressed minimum amount of EGF receptors on their cell surfaces and were refractory to additional EGF in culture. The results indicate that growth factors and their receptors are responsible for tumorigenicity in nude mice.     

Assessment of coronary hemodynamics and myocardial perfusion in patients with syndrome X by digital subtraction angiography and 201Tl-myocardial scintigraphy]

To evaluate coronary hemodynamics and myocardial perfusion, left coronary digital subtraction angiography (DSA) and Tl-201 myocardial scintigraphy were performed in patients with syndrome X. The coronary circulation time (CCT) was significantly prolonged after the injection of isosorbide dinitrate and contrast medium i.c. Apical T1/2 was also prolonged on ergonovine malate provocation test. We suspected that the vascular response of the coronary peripheral artery was impaired, and microvascular spasm probably existed in patients with syndrome X. The prevalence of abnormal myocardial perfusion defect on exercise Tl-201 SPECT in syndrome X was very high, and coronary hemodynamics was significantly disturbed in the group of syndrome X with abnormal Tl-201 SPECT. Tl-201 lung/heart count ratio significantly increased in syndrome X on treadmill test. Because of this, exercise induced left ventricular dysfunction was suspected. We concluded that the main pathophysiological finding of impaired coronary circulation in syndrome X was microvascular spasm.     

In vitro histoculture of human tumors with fluorescent dye end-points measured by confocal microscopy: high correlation of in vitro and in vivo chemosensitivity.

Tumors from 40 patients and 7 established human xenograft tumor lines were grown in three-dimensional histoculture. A Viable-Cell-Index (VCI) based on fluorescent dyes and Growth Fraction Index (GFI) based on [3H]thymidine incorporation were measured by confocal microscopy and histological autoradiography, respectively, after treatment with cytotoxic agents. Chemosensitivity in vitro with the two methods was correlated with chemosensitivity of the same set of human xenografted tumor lines grown in nude mice. The percent accuracy of in vitro to in vivo correlation with VCI (73%) was higher than GFI (63%). The number of false positives with VCI was 12.1% (4/33), and with GFI was 31.3% (10/32). The results thus indicated that in vitro histoculture with fluorescent vital-dye end-points to measure cell viability is of potential use to determine tumor chemosensitivity.     

Intracapsular tonsillar reduction (partial tonsillectomy): reviving a historical procedure for obstructive sleep disordered breathing in children.

OBJECTIVE: We sought to reintroduce a historical procedure-intracapsular tonsillar reduction (partial tonsillectomy or tonsillotomy)-for tonsillar hypertrophy causing obstructive sleep disordered breathing (OSDB) in children, as well as to determine whether partial tonsillectomy, compared with conventional (total) tonsillectomy when performed by more than one surgeon, is equally effective for the relief of OSDB while resulting in less pain and more rapid recovery. STUDY DESIGN: We conducted a retrospective case series at a tertiary children's hospital. The charts of children who underwent partial tonsillectomy and total tonsillectomy (1998 through 2002) for postoperative complications were reviewed. The caregivers were surveyed to assess postoperative pain, rapidity of recovery, and effectiveness of surgery for relieving symptoms of OSDB. RESULTS: Two hundred forty-three children underwent partial tonsillectomy and 107 children underwent total tonsillectomy. There were no significant differences in immediate and delayed complications between the groups. Both operations were equally effective in relieving OSDB. Children who had partial tonsillectomy had significantly less postoperative pain and significantly more rapid recovery. CONCLUSION: Intracapsular tonsillar reduction with an endoscopic microdebrider relieves OSDB as effectively as conventional tonsillectomy, but results in less postoperative pain and a more rapid recovery.