Influence of filtering on the effective concentration and sterility of a 2% cyclosporine ophthalmic solution: a quality improvement perspective.

BACKGROUND: Pharmaceutical companies do not sell formulations for all diseases; thus, healthcare workers have to treat some diseases by concocting in-hospital preparations. An example is the high-concentration 2% cyclosporine A (CyA) ophthalmic solution. Utilizing a filter in sterility operations is a general practice for concocting in-hospital preparations, as is the case for preparing a 2% CyA ophthalmic solution. However, whether filtering is appropriate concerning the active ingredient content and bacterial contamination according to the post-preparing quality control of a 2% CyA ophthalmic solution is yet to be verified. METHODS: We conducted particle size, preparation concentration, and bacterial contamination studies to clarify aforementioned questions. First, we measured the particle size of CyA through a laser diffraction particle size distribution. Next, we measured the concentration after preparation with or without a 0.45-µm filter operation using an electrochemiluminescence immunoassay. Finally, bacterial contamination tests were conducted using an automated blood culture system to prepare a 2% CyA ophthalmic solution without a 0.45 µm filtering. Regarding the pore size of the filter in this study, it was set to 0.45 µm with reference to the book (the 6th edition) with recipes for the preparation of in-hospital preparations edited by the Japanese Society of Hospital Pharmacists. RESULTS: CyA had various particle sizes; approximately 30% of the total particles exceeded 0.45 µm. The mean ± standard deviation of filtered and non-filtered CyA concentrations in ophthalmic solutions were 346.51 ± 170.76 and 499.74 ± 76.95ng/mL, respectively (p = 0.011). Regarding bacterial contamination tests, aerobes and anaerobes microorganisms were not detected in 14 days of culture. CONCLUSIONS: Due to the results of this study, the concentration of CyA may be reduced by using a 0.45-µm filter during the preparation of CyA ophthalmic solutions, and furthermore that the use of a 0.45-µm filter may not contribute to sterility when preparing CyA ophthalmic solutions.

Disproportionality Analysis on Hypothyroidism With Roxadustat Using the Japanese Adverse Drug Event Database.

Hypoxia-inducible factor prolyl-hydroxylase inhibitor (HIF-PHI) is a novel agent for the treatment of renal anemia. HIF-PHI increases endogenous erythropoietin production by inhibiting the degradation of an erythropoietin transcription factor. Although beneficial effects are expected from HIF-PHI, its novel mechanism raises concerns regarding the risk of potential adverse events. The cases of hypothyroidism, which had not been reported in clinical trials, were reported after the administration of roxadustat in a real-world setting. However, the effects of HIF-PHIs on thyroid function have not yet been fully evaluated. This study aimed to assess the clinical impact of HIF-PHIs on thyroid function using the Japanese Adverse Drug Event Report database, a spontaneous reporting system in Japan, because HIF-PHIs were made available in Japan before they were available in other countries. Although a disproportionality signal for hypothyroidism was detected with roxadustat (reporting odds ratio [ROR]:22.1, 95% confidence interval [CI]:18.3-26.7, no signals were detected with another HIF-PHI, daprodustat (ROR:1.3, 95%CI:0.3-5.4), and epoetin beta pegol (ROR:1.2, 95%CI:0.5-2.7). Signals of hypothyroidism due to roxadustat were also detected regardless of age or sex. Approximately 50% of hypothyroidism cases were reported within 50 days of starting roxadustat use. These results indicate that roxadustat use may be related to the development of hypothyroidism. The need for monitoring of thyroid function should be alerted during roxadustat administration regardless of age or sex.

Peripheral Parenteral Nutrition Solutions and Bed Bath Towels as Risk Factors for Nosocomial Peripheral Venous Catheter-related Bloodstream Infection by Bacillus cereus.

In Japan, China, and Singapore, several studies have reported increased incidences of peripheral venous catheter-related bloodstream infection by Bacillus cereus during the summer. Therefore, we hypothesized that bed bathing with a B. cereus-contaminated "clean" towels increases B. cereus contact with the catheter and increases the odds of contaminating the peripheral parenteral nutrition (PPN). We found that 1) professionally laundered "clean" towels used in hospitals have B. cereus (3.3×104 colony forming units (CFUs) / 25cm2), 2) B. cereus is transferable onto the forearms of volunteers by wiping with the towels (n=9), and 3) B. cereus remain detectable (80∼660 CFUs /50cm2) on the forearms of volunteers even with subsequent efforts of disinfection using alcohol wipes. We further confirmed that B. cereus grow robustly (102 CFUs /mL to more than 106 CFUs /mL) within 24hours at 30°C in PPN. Altogether we find that bed bathing with a towel contaminated with B. cereus leads to spore attachments to the skin, and that B. cereus can proliferate at an accelerated rate at 30°C compared to 20°C in PPN. We therefore highly recommend ensuring the use of sterile bed bath towels prior to PPN administration with catheter in patients requiring bed bathing.

Evaluation of the Antiemetic Effect of Premedication Optimized by Pharmaceutical Support in Cisplatin Regimens.

Cisplatin is classified as a drug with high emetic risk; thus, the use of aprepitant or fosaprepitant in addition to a 5-hydroxytryptamine-3 (5-HT3) receptor antagonist and dexamethasone is recommended for antiemetic therapy. Further, hydration is required to prevent renal dysfunction, and the use of magnesium has been proposed as a part of the hydration procedure. When fosaprepitant is chosen for antiemetic therapy because the patient has dysphagia, and magnesium is added to the hydration procedure, there may be an incompatibility between the two drugs that reduces the antiemetic effect. In our hospital, in a former regimen, these two drugs were administered concurrently as premedication for regimens containing cisplatin. We varied the conditions so that in a revised regimen the two drugs did not come into contact due to pharmaceutical support, and we conducted a retrospective study to determine the difference in the antiemetic effect. The observation period was 2 years (from October 2015 to September 2017) for the former regimen group (n = 89) and 2 years (from October 2017 to September 2019) for the revised regimen group (n = 177). Comparison of the former and revised regimen groups revealed sex (p = 0.012); anticancer drug dosage (p = 0.006); and variation of premedication condition (p = 0.043) as factors affected by the revised regimen. Optimization of the premedication regimen was a form of necessary pharmaceutical support to maintain the patient's QOL.

A case of feline gastrointestinal eosinophilic sclerosing fibroplasia.

Feline gastrointestinal eosinophilic sclerosing fibroplasia was diagnosed in an 8-month-old Scottish fold that had a primary gastrointestinal mass involving the stomach, duodenum and mesenteric lymph nodes. Histopathologically, the most characteristic feature of this mass was granulation tissue with eosinophil infiltration and hyperplasia of sclerosing collagen fiber. Immunohistochemically, large spindle-shaped cells were positive for smooth muscle actin and vimentin. This case emphasizes the importance of feline gastrointestinal eosinophilic sclerosing fibroplasia as a differential diagnosis of gastrointestinal neoplastic lesions such as osteosarcoma and mast cell tumor in cats.

Twenty-one proteins up-regulated in human H-ras oncogene transgenic rat pancreas cancers are up-regulated in human pancreas cancer.

OBJECTIVES: We have established rat models of pancreatic ductal adenocarcinoma (PDAC) in which expression of a human H-ras(G12V) or K-ras(G12V) oncogene regulated by the Cre/lox system drives pancreatic carcinogenesis. Pancreatic ductal adenocarcinoma which develops in H-ras(G12V) and K-ras(G12V) transgenic rats is cytogenetically and histopathologically similar to human PDAC. The present study was designed to determine the feasibility of using the commercially available H-ras(G12V) transgenic rat to find diagnostic protein biomarkers for human pancreatic cancer. METHODS: For an animal model to be useful for searching for protein biomarkers for a disease, it is essential that proteins that are up-regulated in the model are also up-regulated in humans. We used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to compare H-ras(G12V) transgenic rat PDAC with surrounding normal pancreas tissue. RESULTS: We identified 30 up-regulated proteins in the H-ras(G12V) transgenic rat PDAC lesions; importantly, 21 human homologs of these 30 rat proteins are up-regulated in human pancreatic cancer patients. CONCLUSIONS: These results indicate that numerous proteins that are up-regulated in H-ras(G12V) transgenic rat PDAC are also up-regulated in human pancreatic cancer; therefore, this rat model can be used to search for diagnostic biomarkers for this disease.

Testicular gonadoblastoma in two pet domestic rabbits (Oryctolagus cuniculus domesticus).

Testicular gonadoblastoma is an uncommon tumor in domestic animals. The current study describes a testicular gonadoblastoma in 2 pet domestic rabbits (Oryctolagus cuniculus domesticus) based on the histomorphological, immunohistochemical, and ultrastructural characteristics of the tumor. The tumor tissue consisted of discrete tubular structures separated by a fibrous stroma. The tubular structures were composed of large round cells similar in appearance to germ cells in the seminiferous tubules, and small spindle cells with oval or elongated nuclei resembling Sertoli cells. The spindle cells showed peculiar structural patterns arranged in a coronal or follicular pattern, often forming Call-Exner bodies like those in an ovarian granulosa cell tumor. One case was concomitant with seminoma. Immunohistochemistry revealed large round cells that were positive for c-kit and placental alkaline phosphatase, while spindle cells were positive for vimentin and Wilms tumor protein. Ultrastructurally, the spherical eosinophilic material (Call-Exner body) consisted of duplicated basal lamina, and sex cord components surrounded a single degenerative cell similar to a germ cell.

Spontaneous iron accumulation in hepatocytes of a 7-week-old female rat.

Spontaneous iron accumulation in hepatocytes was observed in a 7-week-old female Han Wistar GALAS rat. Very fine yellowish brown pigments, which showed a positive reaction with Berlin Blue stain, were apparent in the cytoplasm close to the bile canaliculi, with a diminishing periportal-to-centrilobular gradient. There were also differences in distribution between and within lobes. Transmission electron microscopy revealed cytosolic ferritin and pericanalicular siderosomes in hepatocytes. No degeneration or necrotic changes were observed, and non-hepatocyte cells did not demonstrate any obvious accumulation of iron. There were no abnormalities in the animal other than this finding in the liver.

An instant cell recognition system using a microfabricated coordinate standard chip useful for combinable cell observation with multiple microscopic apparatuses.

Disposable coordinate standard (CS) chips were fabricated by the ejection of melted polystyrene into a metal mold. The CS chip surface was divided into four parts different in height and width. The edge lines of these parts could be recognized as straight lines 2 mum in width in the microscope view and used as the X and Y axes for the culture dish. The CS chip was attached on the bottom of a culture dish outside. Then the dish was set on the microscope stage and moved by means of a motorized automatic stage. The X-Y coordinates of many single-cells in a culture dish were registered, respectively. Once registered, any single-cell could instantly be brought to the center of the microscope view even after displacing the dish from the stage for a while and setting it again on the stage. Therefore, experimenters can easily search any single-cell in any culture dish on any microscope at any time. Such a system is remarkably useful for various modes of single-cell experiment and named "Suguwaculture," which means "instantly" ("sugu" in Japanese) + "recognizable" ("wakaru" in Japanese) + "culture" (during culture).

Semi-quantitative expression and knockdown of a target gene in single-cell mouse embryonic stem cells by high performance microinjection.

Interactions of multiple genes and associated factors are involved in the differentiation and de-differentiation of embryonic stem (ES) cells. Quantitative analysis of these genes and factors is essential for the elucidation of their mechanism. To meet this requirement, we have investigated various experimental conditions for high performance microinjection into mouse ES cells. A speedy and rhythmic operation was found to be important and was accomplished robotically by using a single-cell manipulation technique and XY-address registrable culture dishes. Among many experimental parameters, the tip size of an injection capillary, the pressure condition, and the DNA concentration in the injection capillary were of critical significance. Their optimum values were 0.5-0.8 microm, 0.7 kgf/cm(2) for 30 ms, and 1-100 ng/microl, respectively. Under these conditions, semi-quantitative control of the EGFP gene expression in mouse ES cells and its knockdown was successfully demonstrated.

Guanine nitration in idiopathic pulmonary fibrosis and its implication for carcinogenesis.

RATIONALE: Nitric oxide (NO)-induced nitrative stress of nucleic acids, as evidenced by guanine nitration, appears to be involved in inflammation-induced carcinogenesis. A high incidence of lung cancer in idiopathic pulmonary fibrosis (IPF) is the major reason for poor prognosis in patients with IPF. OBJECTIVES AND METHODS: We immunohistochemically analyzed the formation and localization of 8-nitroguanine in lung tissues from control subjects, patients with IPF, and patients with lung cancer. MAIN RESULTS: Immunohistochemical analysis of control smoker and nonsmoker lungs showed weak immunoreactivity for 8-nitroguanine, mainly in cytoplasm of bronchial epithelial cells. In addition to the bronchial epithelial cells, metaplastic regenerated epithelial cells overlying dense fibrotic lesions in IPF showed strong 8-nitroguanine staining in the cytoplasm. The staining in these metaplastic cells colocalized with staining of inducible and endothelial NO synthases and 8-oxodeoxyguanosine, as evidenced by double-immunostaining analysis. Confocal and immunoelectron microscopy revealed localization of 8-nitroguanine in metaplastic epithelial cytoplasm, mostly in mitochondria. Appreciable 8-nitroguanine immunostaining was also observed in both nuclei and cytoplasm of malignant epithelial cells in squamous cell carcinoma. No significant difference was found in the epithelial 8-nitroguanine formation between control smokers and nonsmokers, but much higher guanine nitration was observed in patients with IPF than in control subjects and patients with lung cancer, via a quantitative immunofluorescence image analysis. CONCLUSIONS: The present study indicates that not only oxidative stress but also nitrative stress induced by NO may participate in the pathogenesis of epithelial cell damage and aberrant regeneration occurring in IPF. Thus, guanine nitration may be a major risk factor for lung cancer development in IPF.

Susceptibilities of p53 knockout and rasH2 transgenic mice to urethane-induced lung carcinogenesis are inherited from their original strains.

In the present study, susceptibility of CB6F1 mice carrying the human prototype c-Ha-ras gene (rasH2 mice) and p53 gene knockout mice (p53 (+/-) mice) to urethane-induced lung carcinogenesis was compared under the same experimental conditions. Both strains were administered 500 ppm urethane in their drinking water for 3 weeks. At week 26, lung adenocarcinomas and adenomas were observed in 53% and 100% of rasH2 mice, respectively, and lung adenomas were observed in 67% of rasH2 littermate (non-Tg) mice. However, lung tumors were not observed in either p53 (+/-) or p53 (+/+) mice. Peliosis hepatis and hepatic hemangiomas were observed in 27% and 67% of p53 (+/-) mice, but only in 6.7% and 6.7% of the rasH2 animals, respectively. Under the same experimental conditions, BALB/c mice, the strain of origin of the rasH2 mice, developed lung adenomas at an incidence of 93%, whereas none of the C57BL/6 original strain for p53 (+/-) mice developed lung tumors. Peliosis hepatis was observed in 40% of the C57BL/6 mice, but not in BALB/c mice; hepatic and splenic hemangiomas were not observed in these animals. These results indicate that organ susceptibility of rasH2 and p53 (+/-) mice is inherited from their strains of origin, the rasH2 and BALB/c lines being much more sensitive to the induction of pulmonary carcinogenesis.

Mechanisms of promotion and progression of preneoplastic lesions in hepatocarcinogenesis by DDT in F344 rats.

Time-related changes in potential factors involved in hepatocarcinogenesis by DDT were investigated in a 4-week and a 2-year feeding studies of p,p'-DDT with F344 rats. In the 4-week study with males at doses of 50, 160, and 500 ppm, cell proliferation and gap junctional intercellular communication (GJIC) were examined after 1, 2, 3, 7, 14, and 28 days. Cell proliferation was enhanced within 3 days at any dose level, but returned to normal after 7 days, whereas GJIC was inhibited throughout the study. In the 2-year study with both sexes at doses of 5, 50, and 500 ppm, cell proliferation, GJIC, enzyme induction, and oxidative stress were investigated after 26, 52, 78, and 104 weeks. Males and females showed an inhibition of GJIC and increases in P450 isozymes (CYP2B1 and CYP3A2) in a dose-dependent manner at all time points, but no significant change in cell proliferation. Lipid peroxide for males at 50 and 500 ppm and 8-hydroxydeoxyguanosine for both sexes at 500 ppm were elevated throughout the study. Histologically, eosinophilic foci and hepatocellular adenomas increased in males at 50 ppm and both sexes at 500 ppm. Hepatocellular carcinomas also developed in males at 500 ppm. These results indicate that DDT may induce eosinophilic foci as a result of oxidative DNA damage and leads them to neoplasms in combination with its mitogenic activity and inhibitory effect on GJIC. Oxidative stress could be a key factor in hepatocarcinogenesis by DDT.