Deep lamellar keratoplasty by deep parenchyma detachment from the corneal limbs.

AIM: To improve the deep lamellar keratoplasty technique. METHOD: For the easy and reliable perfomance of deep lamellar keratoplasty (DLKP), detachment of Descemet's membrane through the corneal limber flap was improved. To expose Descemet's membrane, the parenchyma was detached by hydrodelamination through a sclerocorneal flap made in the corneal limbs. The parenchyma was removed after the pseudochamber between it and Descemet's membrane was maintained with viscoelastic material. The corneal graft was placed with a running suture. 22 eyes were treated. RESULTS: Complete exposure of Descemet's membrane was obtained in 20 of the 22 eyes (91%). The membrane was perforated in five of the 22 eyes (23%) during surgery, and two of the 22 eyes (9%) were converted to penetrating keratoplasty. These two eyes developed keratoconus after acute corneal hydrops. CONCLUSION: Compared with the conventional procedure, this new method provides easy, reliable exposure of Descemet's membrane.

Effects of acetate and butyrate on the expression of leptin and short-form leptin receptor in bovine and rat anterior pituitary cells.

The effects of acetate and butyrate on leptin and leptin receptor (OB-R) expression in bovine and rat anterior pituitary were examined. In bovine tissues, leptin gene expression using RT-PCR was observed in fat and anterior pituitary but not in liver. Isolated anterior pituitary cells cultured in Dulbecco's modified Eagle's medium (DMEM) for 3 days were further cultured for 48 h in DMEM containing 10 mM acetate or butyrate or without any fatty acids as control. Western blot analysis revealed that the abundance of leptin protein was greater in the presence of acetate and butyrate than that for the control culture. Leptin abundance was increased in a dose- and time-dependent manner in bovine anterior pituitary cells. However, leptin expression in rat cells, of which the basal level was much greater than that in ovine cells, was significantly decreased by the culture with butyrate. In addition, we studied the effects of both fatty acids on OB-R mRNA expression using semi-quantitative RT-PCR. The results showed that butyrate significantly decreased the expression in both bovine and rat cells. These findings indicate that acetate and butyrate enhance leptin expression in bovine, but not in rat anterior pituitary cells while butyrate suppresses OB-Ra expression in both rat and bovine pituitaries.

Combined surgery with deep lamellar keratoplasty.

The opportunity to perform two or more ophthalmic operations simultaneously has increased in recent years. In keratoplasty too, the simultaneous operation consisting of cataract extraction and IOL insertion (triple procedure) has become common. Penetrating keratoplasty (PKP) and planned extracapsular cataract extraction (PECCE) or vitrectomy is today often done simultaneously. However, phacoemulsification and aspiration (PEA) or 3 ports vitrectomy has to be performed on patients with corneal opacity due to some corneal diseases. In such cases, we performed the PEA or vitrectomy coupled with deep lamellar keratoplasty (DLK), successfully applied to closed eye surgery with corneal opacity. Simultaneous surgery using DLK, free of postoperative endothelial type of rejection, and PEA, enabling cataract extraction by a small incision, is a surgical technique making use of the advantage of the two types of operation. We performed the DLK + PEA + IOL on 17 cases. In the other hand, simultaneous surgery consisting of DLK and vitrectomy may be taken into consideration. The use of DLK will make it possible to perform surgery on urgent vitreoretinal diseases with corneal opacity and cataract. We performed the DLK + 3 ports vitrectomy on 3 cases. The current study describes the advantages and disadvantages of these types of operation.

EDTA: a promoter of proliferation in human corneal endothelium.

PURPOSE: To determine whether it is possible to induce proliferation in the endothelium of older donor corneas by treatment of the intact monolayer with EDTA. METHODS: Corneas from donors 52 to 75 years of age were obtained from an eye bank and were usually cut in quarters to increase sample size. The effect of EDTA dose (0.02-2.0 mg/ml) and incubation time (6, 30, and 60 minutes) on endothelial cell-cell contacts was evaluated by staining for ZO-1, a cell junction marker. Cell death was tested by a commercial live-dead assay. Corneal pieces were incubated for 0, 24, 48, or 60 hours in culture medium (M-199, 10% fetal bovine serum, 10 ng/ml epidermal growth factor, 20 ng/ml fibroblast growth factor) before EDTA treatment. After treatment, pieces were incubated in the same medium for 24, 48, 72, or 96 hours to permit cell cycle entry. Tissue was fixed, stained for Ki67 (a marker for late G1-phase through the M-phase), and mounted in medium containing propidium iodide to visualize all nuclei. Confocal images were evaluated by computer (Image software; NIH, Bethesda, MD) to count Ki67-positive and propidium iodide-stained cells. RESULTS: EDTA released corneal endothelial cell-cell contacts in a dose- and time-dependent manner. At doses and incubation times tested, EDTA did not induce significant cell death. Preincubation in culture medium for 24 hours was needed for endothelial cells to efficiently initiate proliferation in response to EDTA. The endothelium of corneas incubated in mitogen-containing medium for up to 108 hours without EDTA treatment did not stain for Ki67. EDTA at 2.0 mg/ml for 60 minutes appeared optimal and stimulated 16% to 18% of the cells to proliferate. Ki67-positive mitotic figures were visible 48 hours after exposure to EDTA. Formation of daughter cells was visible after double-staining for Ki67 and ZO-1. CONCLUSIONS: EDTA released cells from contact inhibition and promoted proliferation in corneal endothelium from older donors. The authors hypothesize that corneal endothelium from older individuals divide in situ when exposed to positive growth factors under conditions in which cells have been transiently released from contact inhibition.

Cell cycle kinetics in corneal endothelium from old and young donors.

PURPOSE: To compare cell cycle kinetics in corneal endothelial cells from young and old donors. METHODS: Human corneas were obtained from the eye bank and separated into two groups: young (19 corneas, <30 years of age) and old (40 corneas, >50 years of age). Corneas were cut in quarters, and the endothelium was released from contact inhibition by producing a 2-mm scrape wound. Unwounded endothelium acted as a negative control. Corneal pieces were exposed for 24, 36, 48, 60, 72, and 84 hours to medium containing 10% fetal bovine serum, 20 ng/ml fibroblast growth factor, and 50 mg/ml gentamicin or the same medium supplemented with 10 ng/ml epidermal growth factor (EGF). Tissue was fixed, immunostained for Ki67 (a marker for the late G1-through M-phase) or for 5-bromo-2'-deoxyuridine (BrdU; a marker for the S-phase), and mounted in medium containing propidium iodide (PI) to visualize all nuclei. Confocal images were evaluated using an image analysis program to count Ki67-positive and PI-stained cells and to evaluate cell cycle position. Cells were counted in 15x100 microm2 areas randomly selected from each wound, and the mean was used for subsequent calculations. RESULTS: Human corneal endothelial cells could be reliably scored for their position within the cell cycle using Ki67 staining patterns. In both age groups, cells repopulating the wound area stained positively for Ki67, whereas no Ki67 staining was observed in unwounded areas under any condition tested. Cells from old donors treated with fetal bovine serum and FGF stained positively for Ki67, indicating that these cells were actively cycling. Compared with cells from young donors, old cells entered the cell cycle more slowly (48 versus 36 hours), the peak of Ki67 staining occurred later (72 versus 60 hours), and fewer cells proliferated (23% versus 47%) or exhibited mitotic figures (4% versus 7%). Addition of EGF to the culture medium increased Ki67 staining in both groups, but the effect on old cells was more dramatic. More cells from old donors entered the cell cycle by 36 hours after wounding, the number of proliferating cells increased 1.6-fold, and the relative number of mitotic figures increased 2.5-fold over cells treated in the absence of EGF. CONCLUSIONS: Regardless of donor age, corneal endothelial cells can enter and complete the cell cycle. In the presence of fetal bovine serum and FGF, cells from old donors can proliferate but respond more slowly and to a lesser extent than cells from young donors. EGF added to the medium stimulates cells from old donors to enter the cell cycle faster, increases the relative number of actively cycling cells, and increases the number of cells exhibiting mitotic figures. The resultant hypothesis is that it is possible to stimulate a significant proliferative response in corneal endothelial cells from old individuals. Administration of an optimal combination of stimulatory growth factors is required under conditions in which cells have been transiently released from contact inhibition.

[Usefulness of low dose 5'-DFUR (5'-deoxy-5-fluorouridine) for advanced or recurrent breast cancer].

The usefulness of low dose (600 mg/day) 5'-DFUR for advanced or recurrent breast cancer was evaluated. Eight patients out of 38 showed a partial response (response rate 21.1%), 22 patients had no change, and 8 patients had progressive disease. The response rate was 31.0% in soft tissue and 9.1% in bone, both of which were slight. In both one-shot (200 mg) and continuous (1-4 months) administration of 5'-DFUR (600 mg/day), the concentration of 5'-DFUR of 5-FU in sera reached a plateau from 30 minutes to 2 hours after administration of 5'-DFUR, and disappeared after 3 hours. There were no significant differences in the concentration of 5'-DFUR of 5-FU in sera between one-shot and continuous administration. It is concluded that there was no accumulation of the drug after continuous administration of 5'-DFUR, and that low dose 5'-DFUR is a useful therapy for advanced or recurrent breast cancer.

[Stimulation of corneal endothelial cell proliferation by interleukins, complete mitogens and corneal parenchymal cell-derived factors].

The effects of various cytokines and mitogens on proliferation of rabbit corneal endothelial cells were examined in a serum-free culture. Complete mitogens such as epidermal growth factor (EGF), transforming growth factor alpha (TGF-alpha), and hepatocyte growth factor (HGF) stimulated proliferation of corneal endothelial cells 1.5 to 2 times the control. The effect was dose-dependent up to 10 ng/ml and flattened out at higher concentrations. Interleukins (IL-x) such as IL-1 alpha, IL-1 beta, IL-2, IL-3 or IL-4 by themselves had little or no stimulatory effect on cell proliferation. The combination of IL-1 beta and EGF, or IL-1 beta and TGF-alpha, however, showed a 5- to 6-fold stimulation over the control. IL-1 beta, thus, can be regarded as a co-mitogen that can amplify the stimulatory effect of complete mitogens (EGF and TGF-alpha). The number of corneal endothelial cells increased to 200% when corneal endothelial cells were co-cultured with corneal parenchymal cells. Addition of conditioned medium obtained from the culture of corneal parenchymal cells also increased the number of corneal endothelial cells more than 2-fold. These results suggest that factors which stimulate cell proliferation of corneal endothelial cells are secreted by corneal parenchymal cells.

[Effectiveness of combination hormonal therapies in advanced or recurrent breast cancer--a randomized trial of tamoxifen (TAM), medroxyprogesterone acetate (MPA), their simultaneous combination, and an alternating sequential combination of TAM plus MPA].

The effects of tamoxifen (TAM), medroxyprogesterone acetate (MPA), their simultaneous combination or an alternating sequential combination of TAM plus MPA (TAM-MPA), was evaluated in 52 patients with advanced or recurrent breast cancer in a randomized controlled trial. The response rates of TAM, MPA, TAM & MPA, and TAM-MPA were 13.3%, 36.4%, 7.1% and 25.0%, respectively. All four therapies showed a higher response rate in the patients with soft tissues or with no previous therapy. However, there were no significant differences between the patient's characteristics and response rate among these four therapies. The differences in the frequency of adverse effects among these therapies were not significant, and there were no severe adverse effects with any therapy. In the cross-over trial of TAM and MPA, although 15.4% of patients who failed to show a response to TAM showed an objective response to MPA, no response to TAM was seen in the patients who had failed to show a response to MPA. In conclusion, TAM & MPA was not more effective than TAM or MPA, and TAM-MPA was not more effective than MPA. Adverse effects were not increased with TAM & MPA or TAM-MPA. MPA was useful as a second-line therapy after TAM.

[Clinical studies on lung disease caused by atypical mycobacteria in our hospital--especially in relation to drug susceptibility].

From 1987 to 1991, 27 patients were diagnosed as atypical mycobacteriosis in our hospital. Some strains of M. avium complex were found to be M. avium and M. intracellulare by means of a DNA probe test. 1. Total cases consisted of 14 males (the average age was 66.7 years) and 13 females (65.7 years). 2. M. avium complex was observed in 24 patients (9 cases of M. avium and 5 cases of M. intracellulare): M. kansasii and M. chelonae were found in 2 patients and 1 patient, respectively. 3. The findings of sputum cultures became negative three months after the chemotherapy treatment in 3 out of 25 patients. Two male patients were operated on and cured. Three patients died, and all of them had respiratory infections. 4. To determine the susceptibility of mycobacteria strains isolated from patients to various antimicrobial agents, an investigation was carried out. There were 88 strains of M. tuberculosis, 52 strains of M. avium complex (23 strains of M. avium and 17 strains of M. intracellulare), 3 strains of M. kansasii, 2 strains of M. gordonae and 2 strains of M. chelonae. 5. Strains of M. tuberculosis, M. kansasii and M. gordonae were susceptible to various antituberculous agents and ciprofloxacin. Strains of M. chelonae were susceptible to ciprofloxacin, and one strain was susceptible to streptomycin and ethionamide. 6. The M. avium strains were more susceptible to cycloserine and ciprofloxacin than were the M. intracellulare strains. Conversely, the M. intracellulare strains were more susceptible to ethambutol than were the M. avium strains.

Vector quantization for entropy coding of image subbands.

Vector quantization for entropy coding of image subbands is investigated. Rate distortion curves are computed with mean square error as a distortion criterion. The authors show that full-search entropy-constrained vector quantization of image subbands results in the best performance, but is computationally expensive. Lattice quantizers yield a coding efficiency almost indistinguishable from optimum full-search entropy-constrained vector quantization. Orthogonal lattice quantizers were found to perform almost as well as lattice quantizers derived from dense sphere packings. An optimum bit allocation rule based on a Lagrange multiplier formulation is applied to subband coding. Coding results are shown for a still image.

Imprint immunocytochemistry of estrogen receptors in breast carcinoma.

To clarify the accuracy of immunocytochemical detection of estrogen receptors (ER) in breast carcinomas using cytological materials, imprint specimens from tumor tissue were compared with frozen tissue sections and tumors analyzed by the dextran coated charcoal (DCC) method and enzyme immunoassay (EIA). Out of 50 cases examined by imprint immunocytochemistry, there were 39 ER positive cases (78.0% positivity). The positivity in the imprint materials agreed with that of the DCC in 36 out of 40 cases (85.0%), with 100% sensitivity and 60.0% specificity. The two methods statistically correlated with each other in their positivity and grade (p less than 0.001). The positivity and grades of imprint and frozen immunocytochemistry as well as those of imprint immunocytochemistry and the EIA agreed almost perfectly with each other. As a result of the present study, we concluded that immunocytochemical detection of ER is indeed reliable, as accurate as other procedures. We recommend that aspiration biopsy cytology (ABC) be used for morphological examination and ER immunocytochemistry when adequate materials are available and that imprint materials be used when ABC materials are inadequate and fresh tissue is available at the time of surgery.

[Cooperative study of postoperative long-term adjuvant therapy of breast cancer. First study of a series--comparative study between postoperative tegafur administration and no adjuvant chemotherapy].

A prospective randomized study was performed in 22 institutions from July 1978, to evaluate the efficiency of long-term adjuvant chemotherapy with tegafur alone for primary breast cancer. Five hundred and eighty-seven eligible patients, classified into T1a, T2a, T3a, N0, N1a, N1b, and M0 were divided into two groups: Group A, radical mastectomy receiving no adjuvant chemotherapy; and Group B, radical mastectomy followed by 4 courses of adjuvant chemotherapy, each one consisting of daily administration of tegafur 600 mg p.o. for 8 weeks and 4 weeks rest. At the 66th month of median follow up time, there were no differences between the two groups on 5 year cumulative survival and disease free survival rate. In subgroup patients who have histologically 1-3 axillary involvements, there seemed to be a more meaningful prolongation of the disease-free interval in group B than in group A (logrank test; p = 0.11 generalized Wilcoxon test; p = 0.13).

[Practical plate-thermography in the diagnosis of breast cancer].

Described of diagnostic practices of Plate-thermography in breast diseases Described of Takahashi's classification of Plate-thermography referring to both benign and malignant clinical cases. Of 63 cases of breast cancer and 117 cases of benign diseases, the rate of correct diagnosis for cancer was 81% and those of benign disease was 80.1%.

Difference in extractability of estradiol- and tamoxifen-receptor complex in the nuclei from MCF-7 cells with Nonidet P-40.

The extraction of [3H]estradiol- and [3H]tamoxifen-receptor complex in the nuclei from MCF-7 cells with the nonionic detergent Nonidet P-40 has been studied. We found that there is a striking difference in the extractability of estradiol- and tamoxifen-receptor complex from nuclei with 0.5% Nonidet P-40. The nuclear bound estradiol-receptor complex is scarcely extractable with Nonidet P-40. In contrast, almost all of the nuclear bound tamoxifen-receptor complex is extractable. The nuclear [3H]tamoxifen-receptor complex extracted in the presence of Nonidet P-40 sediments in two peaks at 7 S and 5 S. The latter sedimentation rate is the same with that of the nuclear [3H]tamoxifen-receptor complex extracted with 0.4 M KCl. The nuclear [3H]estradiol-receptor complex extracted with 0.4 M KCl sediments at 4 S. The results suggest that interaction of tamoxifen-receptor complex with chromatin is different from that of estradiol-receptor complex.