Tissue engineering of corneal stroma with rabbit fibroblast precursors and gelatin hydrogels

PURPOSE: To isolate fibroblast precursors from rabbit corneal stroma using a sphere-forming assay, to engineer corneal stroma with the precursors and gelatin, and to establish the therapeutic application of precursors in a rabbit corneal stroma. METHODS: In the in vitro study, a sphere-forming assay was performed to produce precursors from rabbit corneal stroma. Corneal stroma was engineered by cultivating precursors in porous gelatin for one week. In the in vivo study, the engineered corneal stromal sheet with precursors (precursor/gelatin group) or with fibroblasts (fibroblast /gelatin group) or without cells (gelatin group) was transplanted to a pocket of rabbit corneal stroma. Gene expression and extracellular matrix production were examined immunohistochemically in each group one week and four weeks after surgery. RESULTS: In the in vitro study, cells in the spheres were BrdU-positive, and their progeny were keratocan-positive. The study also showed that the corneas transplanted with a porous gelatin sheet did not show any opacity four weeks after transplantation in any group. In the gelatin sheet of the precursor/gelatin group, a more intense expression of type I collagen was observed relative to the other two groups four weeks after the surgery. CONCLUSIONS: Our findings demonstrate that the transplantation of fibroblast precursors combined with gelatin hydrogel into the corneal stroma is a possible treatment strategy for corneal stromal regeneration

Effects of cellooligosaccharide or a combination of cellooligosaccharide and live Clostridium butyricum culture on performance and intestinal ecology in Holstein calves fed milk or milk replacer

The effects of oral administration of a prebiotic (cellooligosaccharide [CE]) and a combination of a probiotic (a commercial Clostridium butyricum strain) and prebiotics (referred to as symbiotics [SB]) on performance and intestinal ecology in Holstein calves fed milk replacer (MR) or whole milk were evaluated. Forty female calves (experiment 1) and 14 male and female calves (experiment 2) were used in this study. Calves were fed MR (experiment 1) or whole milk (experiment 2) necessary for daily weight gain of 0.3 kg based on birth weight in two daily feedings and weaned at 46 days. Calves were divided into a CE feeding group, SB feeding group (only in experiment 1), and control group. The CE and SB groups were fed CE at 5 g/day before weaning and 10 g/day postweaning. Only the SB group received 108 colony-forming units (CFU) of C butyricum culture per day. Commercial calf starter was offered for ad libitum intake. Health and feed intake of the animals were monitored daily, and body weight was measured weekly. Fecal samples were analyzed for determination of bacterial community composition by an RNA-based method (sequence-specific SSU rRNA cleavage method) and for organic acid profiling. In 49-day experiments, feed intake, daily gain, and occurrence of diarrhea of the calves were unaffected by either CE supplementation or SB supplementation, and all calves were healthy during each experiment. The fecal bacterial community compositions and the organic acid profiles were not different among groups in experiment 1. In experiment 2, the level of the Clostridium coccoides-Eubacterium rectale group was higher in the feces of CE group than controls at 4 weeks of age and fecal butyric acid concentration was higher (8.0 vs. 12.2 [mmol/kg feces], P <0.05) at that time. There were no differences in prebiotic bacteria (the genera Lactobacillus and Bifidobacterium) between groups at this time point. These results suggested that CE and C. butyricum supplementation have less effect on the performance of healthy calves fed MR. However, prebiotic supplementation seems effective for modulation of the intestinal bacterial community of calves when administered with whole milk.ArticleLIVESTOCK SCIENCE. 153(1-3):88-93 (2013)journal articl

Cytopathic effects and local immune responses in repeated neoadjuvant HSV-tk + ganciclovir gene therapy for prostate cancer

ObjectiveCytopathic effects and local immune response were analyzed histologically in prostatic carcinoma (PCa) with in situ herpes simplex virus-thymidine kinase (HSV-tk)/ganciclovir (GCV) gene therapy (GT). MethodsFour high-risk PCa patients who received HSV-tk/GCV GT were investigated. After two cycles of intraprostatic injection of HSV-tk and administration of GCV, radical prostatectomy was performed. Formalin-fixed, paraffin-embedded sections were evaluated using immunohistochemistry. PCa with hormone therapy (HT, n = 3) or without neoadjuvant therapy (NT, n = 4) that were equivalent in terms of risk were also examined as reference. Immunoreactively-positive cells were counted in at least three areas in cancer tissue. Labeling indices (LI) were calculated as percentage values. ResultsssDNA LI in GT increased, indicating apoptosis, as well as tumor-infiltrating lymphocytes and CD68-positive macrophages, compared with their biopsies. GT cases showed significantly higher numbers of ssDNA LI, CD4/CD8-positive T cells and CD68-positive macrophages including M1/M2 macrophages than HT or NT cases. However, there was no significant difference in CD20-positive B cells among the types of case. There were strong correlations between CD8+ T cells and CD68+ macrophages (ρ = 0.656, p < 0.0001) as well as CD4+ T cells and CD20+ B cells (ρ = 0.644, p < 0.0001) in PCa with GT. ConclusionsEnhanced cytopathic effect and local immune response were might be indicated in PCa patients with HSV-tk/GCV gene therapy.Cytopathic effects and local immune response were analyzed histologically in prostatic carcinoma (PCa) with in situ herpes simplex virus-thymidine kinase (HSV-tk)/ganciclovir (GCV) gene therapy (GT..

A Phthalimide Derivative That Inhibits Centrosomal Clustering Is Effective on Multiple Myeloma

Despite the introduction of newly developed drugs such as lenalidomide and bortezomib, patients with multiple myeloma are still difficult to treat and have a poor prognosis. In order to find novel drugs that are effective for multiple myeloma, we tested the antitumor activity of 29 phthalimide derivatives against several multiple myeloma cell lines. Among these derivatives, 2-(2,6-diisopropylphenyl)-5-amino-1H-isoindole-1,3- dione (TC11) was found to be a potent inhibitor of tumor cell proliferation and an inducer of apoptosis via activation of caspase-3, 8 and 9. This compound also showed in vivo activity against multiple myeloma cell line KMS34 tumor xenografts in ICR/SCID mice. By means of mRNA display selection on a microfluidic chip, the target protein of TC11 was identified as nucleophosmin 1 (NPM). Binding of TC11 and NPM monomer was confirmed by surface plasmon resonance. Immunofluorescence and NPM knockdown studies in HeLa cells suggested that TC11 inhibits centrosomal clustering by inhibiting the centrosomal-regulatory function of NPM, thereby inducing multipolar mitotic cells, which undergo apoptosis. NPM may become a novel target for development of antitumor drugs active against multiple myeloma

A Phase I Trial of 100mg/m2 Docetaxel in Patients with Advanced or Recurrent Breast Cancer

Docetaxel is a standard treatment for patients with advanced or recurrent breast cancer. The recommended dose is 60 to 100 mg/m2. Previous study have shown that the tumor response rates of patients who received docetaxel monotherapy at doses of 60, 75, and 100 mg/m2 were 22.1% , 23.3% , and 36.0% , respectively, and there was a significant relationship between the dose and response. In Europe and the United States, docetaxel is approved at a dose of 100 mg/m2, and Japanese guidelines also recommend a dose of 100 mg/m2. However, the approved dose in Japan is up to 75 mg/m2. We have launched a phase I trial evaluating 100 mg/m2 docetaxel in patients with advanced or relapsed breast cancer. The major eligibility criteria are as follows: age &ge;20 years, pathologically diagnosed breast cancer, recurrent or advanced breast cancer, a good performance status, and HER2 [human epidermal growth factor receptor 2] negative. The primary endpoint is demonstrated safety of 100 mg/m2 docetaxel. This study will clarify whether 100mg/m2 docetaxel can be administrated safely in Japanese patients with advanced or recurrent breast cancer

Topical insulin-like growth factor 1 treatment using gelatin hydrogels for glucocorticoid-resistant sudden sensorineural hearing loss: a prospective clinical trial

<p>Abstract</p> <p>Background</p> <p>Sudden sensorineural hearing loss (SSHL) is a common condition in which patients lose the hearing in one ear within 3 days. Systemic glucocorticoid treatments have been used as standard therapy for SSHL; however, about 20% of patients do not respond. We tested the safety and efficacy of topical insulin-like growth factor 1 (IGF1) application using gelatin hydrogels as a treatment for SSHL.</p> <p>Methods</p> <p>Patients with SSHL that showed no recovery to systemic glucocorticoid administration were recruited. We applied gelatin hydrogels, impregnated with recombinant human IGF1, into the middle ear. The primary outcome measure was the proportion of patients showing hearing improvement 12 weeks after the test treatment. The secondary outcome measures were the proportion of patients showing improvement at 24 weeks and the incidence of adverse events. The null hypothesis was that 33% of patients would show hearing improvement, as was reported for a historical control after hyperbaric oxygen therapy.</p> <p>Results</p> <p>In total, 25 patients received the test treatment at a median of 23 days (range 15-32) after the onset of SSHL, between 2007 and 2009. At 12 weeks after the test treatment, 48% (95% CI 28% to 69%; <it>P </it>= 0.086) of patients showed hearing improvement, and the proportion increased to 56% (95% CI 35% to 76%; <it>P </it>= 0.015) at 24 weeks. No serious adverse events were observed.</p> <p>Conclusions</p> <p>Topical IGF1 application using gelatin hydrogels is well tolerated and may be efficacious for hearing recovery in patients with SSHL that is resistant to systemic glucocorticoids.</p

Effects of cellooligosaccharide or a combination of cellooligosaccharide and live Clostridium butyricum culture on performance and intestinal ecology in Holstein calves fed milk or milk replacer

The effects of oral administration of a prebiotic (cellooligosaccharide [CE]) and a combination of a probiotic (a commercial Clostridium butyricum strain) and prebiotics (referred to as symbiotics [SB]) on performance and intestinal ecology in Holstein calves fed milk replacer (MR) or whole milk were evaluated. Forty female calves (experiment 1) and 14 male and female calves (experiment 2) were used in this study. Calves were fed MR (experiment 1) or whole milk (experiment 2) necessary for daily weight gain of 0.3 kg based on birth weight in two daily feedings and weaned at 46 days. Calves were divided into a CE feeding group, SB feeding group (only in experiment 1), and control group. The CE and SB groups were fed CE at 5 g/day before weaning and 10 g/day postweaning. Only the SB group received 108 colony-forming units (CFU) of C butyricum culture per day. Commercial calf starter was offered for ad libitum intake. Health and feed intake of the animals were monitored daily, and body weight was measured weekly. Fecal samples were analyzed for determination of bacterial community composition by an RNA-based method (sequence-specific SSU rRNA cleavage method) and for organic acid profiling. In 49-day experiments, feed intake, daily gain, and occurrence of diarrhea of the calves were unaffected by either CE supplementation or SB supplementation, and all calves were healthy during each experiment. The fecal bacterial community compositions and the organic acid profiles were not different among groups in experiment 1. In experiment 2, the level of the Clostridium coccoides-Eubacterium rectale group was higher in the feces of CE group than controls at 4 weeks of age and fecal butyric acid concentration was higher (8.0 vs. 12.2 [mmol/kg feces], P <0.05) at that time. There were no differences in prebiotic bacteria (the genera Lactobacillus and Bifidobacterium) between groups at this time point. These results suggested that CE and C. butyricum supplementation have less effect on the performance of healthy calves fed MR. However, prebiotic supplementation seems effective for modulation of the intestinal bacterial community of calves when administered with whole milk.ArticleLIVESTOCK SCIENCE. 153(1-3):88-93 (2013)journal articl