Investigation into the Physiological State of Heat Stressed Escherichia coli Used in the Evaluation Testing of an Intrinsic Fluorescence-Based RMM.

Rapid microbiological methods (RMMs) have been used as novel quality control technologies in industry. The ability of RMMs to detect stressed bacteria, in particular, is of continued interest due to the limitations of the conventional method in stressed bacteria detection. Accordingly, there is a need to better characterize an RMM's ability to detect stressed microorganisms. Previously we reported on the detection ability of an intrinsic fluorescence-based RMM using a 50% injured (determined based on colony-forming ability) bacterial cell group after heat treatment at 55°C for 8 min. In this study, we added further information about the physiological state of the heat treated Escherichia coli, besides proliferation ability, by investigating respiratory activity using CTC fluorescent staining and expression of DnaK, a heat shock protein. It was found that 89% of cells (control 96%) retained respiratory activity, but only 20% (control 41%) retained proliferation ability after heat treatment. The difference between the percentage of cells with respiratory activity versus that of cells still capable of proliferation further supports the existence of viable but non-culturable stressed cells in the test sample. Also, we suggest such analysis would be one approach to confirming the use of stressed as opposed to dead cells when evaluating an RMM's ability to detect stressed microorganisms.

Evaluation of Navigated Laser Photocoagulation (Navilas 577+) for the Treatment of Refractory Diabetic Macular Edema.

PURPOSE: To evaluate navigated laser photocoagulation for the treatment of refractory diabetic macular edema (DME). METHODS: Retrospective study of 25 eyes (21 patients) treated with Navilas 577+ focal laser system. Best-corrected visual acuity (BCVA) and spectral-domain optical coherence tomography (OCT) parameters were measured at baseline, 1, 3, and 6 months, and final visit. RESULTS: The mean follow-up period was 12.8 ± 2.4 (7-16 months). All subjects had history of previous treatment which was injection of triamcinolone acetonide or antivascular endothelial growth factor (VEGF) agents. The navigated laser photocoagulation was delivered to the microaneurysms on indocyanine green angiography (ICGA) in 21 of 25 eyes (84%), fluorescein angiography (FA) guided in 3 eyes, and OCT angiography guided in 1 eye. After initial navigated laser treatment, 16 of 25 eyes (64%) were needed additional navigated laser photocoagulation, injection of triamcinolone acetonide, and/or injection of VEGF agents. Although median BCVA remained stable, the central retinal thickness and macular volume were significantly decreased over 6 months (p < 0.05). All patients were treated without complications. CONCLUSIONS: Focal photocoagulation using Navilas 577+ showed to be effective in treating DME with improvement in macular edema on OCT over 6 months. Navilas 577+ was beneficial to perform navigated laser photocoagulation based on three modalities (ICGA, FA, and OCT angiography).

Suppression of Retinal Neovascularization by Anti-CCR3 Treatment in an Oxygen-Induced Retinopathy Model in Mice.

PURPOSE: To investigate the association between retinal neovascularization and the CC chemokine receptor-3 (CCR3) in a mouse model of oxygen-induced retinopathy (OIR). METHODS: An OIR model in C57BL/6J mice was used as a retinal neovascularization model. An enzyme-linked immunosorbent assay was performed to evaluate the chronological change in vascular endothelial growth factor A (VEGF-A) and eotaxin expressions. CCR3 and VEGF subtype expression in the retina was examined using real-time RT-PCR, and CCR3, eotaxin, VEGF-A, and CD31 expression was examined immunohistochemically. A CCR3 neutralizing antibody (Ab) was injected into the vitreous humor on both postnatal days 12 (P12) and 14 (P14). Retinal neovascularizations were quantified by measurement of the percentages of neovascular area. RESULTS: The mean eotaxin and VEGF-A protein level was significantly downregulated at P10 and P12 and was significantly upregulated at P14 and P17 (p < 0.05). CCR3 mRNA expression was significantly upregulated at P12 (p < 0.05). VEGF164 mRNA expression was significantly upregulated at P14 (p < 0.05). The areas of vaso-obliteration and neovascularization were significantly suppressed in anti-CCR3 Ab-treated eyes (p < 0.05). Anti-CCR3 Ab treatment suppressed VEGF and eotaxin but not monocyte chemoattractant protein-1. And VEGF 164 mRNA but not VEGF120 mRNA was suppressed by anti-CCR3 Ab treatment. CONCLUSIONS: The present data suggest that anti-CCR3 treatment can suppress retinal neovascularization. Anti-CCR3 treatment may have potential as a new therapy for retinopathies with retinal neovascularization such as diabetic retinopathy and retinopathy of prematurity.

Evaluation of peripheral fundus autofluorescence in eyes with wet age-related macular degeneration.

PURPOSE: We aimed to evaluate the prevalence of abnormal peripheral fundus autofluorescence (FAF) in wet age-related macular degeneration (AMD) using wide-field imaging instrument. PATIENTS AND METHODS: A retrospective, case-controlled study involving 66 eyes of 46 Japanese wet AMD patients and 32 eyes of 20 control patients was performed. Wide-field FAF images were obtained for typical AMD (37 eyes/28 patients), polypoidal choroidal vasculopathy (PCV) (22 eyes/20 patients), and retinal angiomatous proliferation (RAP) (seven eyes/four patients). Two masked ophthalmologists independently graded the images for mottled, granular, and nummular patterns. Main outcome measures were abnormal peripheral FAF frequencies and relative risks by disease subgroups and treatments. RESULTS: Abnormal peripheral FAF patterns were found in 51.5% of wet AMD eyes compared with 18.8% of control eyes (P<0.001). Mottled, granular, and nummular patterns were found in 45.5%, 31.8%, and 16.7%, respectively, of wet AMD eyes. Each disease subgroup (typical AMD, 54.1%; PCV, 36.4%; and RAP, 85.7%) showed significantly higher frequencies of peripheral FAF (P<0.001, P=0.03, and P<0.001, respectively) than control eyes (18.8%). There were no significant differences (P=0.76) between the frequencies in untreated and treated eyes. CONCLUSION: Eyes of Japanese wet AMD patients had a higher abnormal FAF prevalence compared with control eyes. Among the three disease subtypes, abnormal patterns were least prevalent in PCV eyes.

Tissue Plasminogen Activator as an Antiangiogenic Agent in Experimental Laser-Induced Choroidal Neovascularization in Mice.

PURPOSE: We investigate the antiangiogenic efficacy of tissue plasminogen activator (tPA) on experimental laser-induced choroidal neovascularization (CNV) in mice. METHODS: After CNV was induced by laser photocoagulation in 92 C57BL/6J wild-type mice, tPA (4 or 40 international units [IU]/µl) or PBS was injected intravitreally immediately after laser injury. Fluorescein angiography was performed on day 7 to grade CNV leakage. The CNV volume was measured by confocal microscopy in eyes enucleated 7 days after laser injury. Immunohistochemical studies were performed 3 days after laser injury to evaluate fibrin/fibrinogen and CD31 expression. The possible adverse effects of tPA were assessed by electroretinography (ERG) and histology on day 7. RESULTS: Intravitreal administration of tPA significantly suppressed CNV leakage and CNV volume in a dose-dependent manner (P < 0.01). Intravitreal injection of tPA suppressed fibrin/fibrinogen and CD31 expression in laser-induced lesions. Histologic examination and ERG showed no evidence of retinal toxicity in eyes injected with tPA. CONCLUSIONS: Intravitreal injection of tPA suppressed fibrin/fibrinogen expression and laser-induced CNV. The current results suggested that tPA may be a potential therapeutic adjuvant for treating CNV.

New Insights Into Microaneurysms in the Deep Capillary Plexus Detected by Optical Coherence Tomography Angiography in Diabetic Macular Edema.

PURPOSE: To study the association between the distributions of microaneurysms detected by en face optical coherence tomography angiography (OCTA) and diabetic macular edema (DME). METHODS: The study design was a retrospective chart review of 27 patients (33 eyes) with DME. The eyes were scanned using OCTA (6 × 6 mm) and spectral-domain (SD) OCT macular cube. Each of the images of the capillary plexus was overlaid onto the image of the topographic map, and the densities of the microaneurysms were measured by ImageJ software. The association between the distribution of microaneurysms and macular edema was evaluated. RESULTS: For microaneurysms in areas with and without edema, 77.3 ± 8.1% of these microaneurysms were located in the deep capillary plexuses (DCP). However, in areas of edema where the retinal thickness was more than 400 µm, 91.3 ± 9.1% of the microaneurysms were found in the DCP. This difference was statistically significant (P < 0.001). In the macular edema area, there was a significantly higher density of microaneurysms in the DCP compared to the superficial capillary plexuses (1.71/mm2 vs. 0.17/mm2, P < 0.001). There was also a significant correlation between the macular volume and the density of microaneurysms in the DCP in edema (r = 0.63, P < 0.001). CONCLUSIONS: Our study demonstrated a high proportion of microaneurysms in the DCP, as well as a novel association between the distributions of microaneurysms detected by OCTA and DME. Results also indicated that microaneurysms located in the DCP contribute to the pathogenesis of DME.

Investigation of the detection ability of an intrinsic fluorescence-based bioaerosol detection system for heat-stressed bacteria.

A number of rapid microbiological methods capable of aerosol-based microbial detection are quickly emerging for use in the pharmaceutical and food markets. A subset of these technologies utilizes intrinsic microbial fluorescence as the basis for bioaerosol detection. This fundamental method of detection is relatively new to the pharmaceutical and food industries, which rely on traditional culture-based methods implemented decades ago to gain an understanding of their manufacturing environments. When combined with real time and continuous assessment, intrinsic fluorescence-based detection provides a new level of information and monitoring in these environments. One aspect of this monitoring relates to the detection of stressed micro-organisms. Bacteria found in pharmaceutical and food manufacturing environments can be in a stressed state due to heat, UV, or chemical exposure, desiccation, and so forth. As a result, the ability of an environmental monitoring system to detect stressed microbes is of particular interest. A commercially available, intrinsic fluorescence-based bioaerosol detection RMM was utilized in this study to determine the ability of such systems in the detection of heat-stressed microorganisms. An assessment of culturability and growth delay in control and heat-stressed samples was performed to confirm stress. Furthermore, the performance of the intrinsic fluorescence-based bioaerosol detection systems were compared to the SAS Super 100, MAS-100 NT, and SMA air samplers in the detection of heat-stressed Escherichia coli, Staphylococcus epidermidis, and Bacillus atrophaeus spores. These bacteria were selected because they are industry-relevant organisms, commonly found in various manufacturing environments, that represent a Gram-positive, Gram-negative, and spore-forming bacteria, respectively. It was found that the intrinsic fluorescence-based bioaerosol detection systems can detect heat-stressed microorganisms, including those that are not detected by the traditional culture-based method due to the inability of the stressed microbes to form colony-forming units. LAY ABSTRACT: Rapid microbiological methods capable of aerosol-based microbial detection are emerging for use in pharmaceutical and food markets. A subset of these technologies utilizes intrinsic microbial fluorescence as the basis for bioaerosol detection, a method relative new to the pharmaceutical and food industries, which rely on traditional culture-based methods. Bacteria found in such environments can be in a stressed state due to heat, UV, or chemical exposure, desiccation, and so forth. As a result, the ability of an environmental monitoring system to detect stressed microbes is of particular interest. A commercially available, intrinsic fluorescence-based bioaerosol detection RMM was utilized in this study to determine the ability of such systems in the detection of heat-stressed microorganisms. An assessment of culturability and growth delay in control and heat-stressed samples was performed to confirm stress. Furthermore, the performance of the intrinsic fluorescence-based bioaerosol detection systems were compared to the SAS Super 100, MAS-100 NT and SMA air samplers in the detection of heat-stressed Escherichia coli, Staphylococcus epidermidis, and Bacillus atrophaeus spores. These bacteria were selected because they are industry-relevant organisms, commonly found in various manufacturing environments, that represent a Gram-positive, Gram-negative, and spore forming bacteria, respectively. It was found that the intrinsic fluorescence-based bioaerosol detection systems can detect heat-stressed microorganisms, including those that are not detected by the traditional culture-based method due to the inability of the stressed microbes to form colony-forming units.

Quantitative comparison of the autofluorescence of bacteria and polystyrene microspheres under violet wavelength excitation for verification of fluorescence-based bioaerosol detector results.

The autofluorescence intensity of bacteria and fungal spores was quantified by fluorescence microscopy in order to obtain the information for evaluating fluorescence-based bioaerosol detectors and was comparable to that of some types of polystyrene microspheres (PSMs). Although the intensity for microbes was distributed across a wide range over an order of magnitude in gray scale, it was in the intensity range of certain PSMs. Furthermore, some of those bacteria and PSMs were aerosolized in a test chamber and the fluorescence intensity was measured with a bioaerosol detector. Although there was a slight difference in the order of intensity from the results obtained by fluorescence microscopy, the fluorescence-based bioaerosol detector showed the intensity was in a comparable range.

Mei4p coordinates the onset of meiosis I by regulating cdc25+ in fission yeast.

The kinase Cdc2p is a central regulator of entry into and progression through nuclear division during mitosis and meiosis in eukaryotes. Cdc2p is activated at the onset of mitosis by dephosphorylation on tyrosine-15, the phosphorylation status of which is determined mainly by the kinase Wee1p and the phosphatase Cdc25p. In fission yeast, the forkhead-type transcription factor Mei4p is required for expression of many genes during meiosis, with mei4 mutant cells arresting before meiosis I. The mechanism of cell cycle arrest in mei4 cells has remained unknown, however. We now show that cdc25(+) is an important target of Mei4p in control of entry into meiosis I. Forced dephosphorylation of Cdc2p on tyrosine-15 thus induced meiosis I in mei4 mutant cells without a delay, although no spores were formed. We propose that Mei4p acts as a rate-limiting regulator of meiosis I by activating cdc25(+) transcription in coordination with other meiotic events.

[Simultaneous renal cell carcinoma and urothelial carcinoma in ipsilateral kidney, report of two cases].

Two cases of simultaneously developed renal tumors (renal cell carcinoma and renal pelvic urothelial carcinoma) in the ipsilateral kidney are reported. These underwent radical surgery under the diagnosis of renal cell carcinoma or renal pelvic urothelial carcinoma, respectively. The risk of incorrect preoperative diagnosis is discussed in terms of insufficient surgical margins. The liberal use of frozen section diagnosis and more radical resection if applicable may be the key to solve such dilemmas.

[Primary, nonviral, hepatocellular carcinoma in patients with prostate cancer treated by hormone therapy: 2 case reports].

We studied two cases of primary, hepatocellular carcinoma (HCC) that occurred following hormone therapy (estrogen therapy in one case and total androgen blockade therapy in another) for stage D2 prostate cancer. Prostate cancer is considered to be hormone-dependent, and androgens appear to be important hormonal factors. However, hepatocellular carcinoma has been shown to have both estrogen and androgen receptors, suggesting that this may be dependent on estrogen or androgen. Reported here are two unique cases of hepatocellular carcinoma in patients with prostate cancer; the pathogenesis of HCC in these patients was suspected to be related to diethylstilbestrol (DES) therapy and antiandrogen therapy for their prostate cancer.

[Urologic area].

In this review, the clinical experience of a three-dimensional(3D) image at urologic area are described. There are some methods for reconstructive 3D image of Multi-slice CT. Maximum intensity projection (MIP) is usefulness for detection of vascular anatomy. Multi-planar reconstruction (MPR) is usefulness for relation with circumference organs. Therefore the opportunities of angiography decreased. It seems that an applied range of a 3D image spreads more in future.

[Prostatic cancer in a young adult: a report of 2 cases].

Patients younger than 45 years with prostate cancer are rare. Between 1999 and 2002, we studied two cases of prostate cancer in men aged under 45 years. Case 1; a 45-year-old man admitted with the chief complaint of urination disorder. Serum level of prostate-specific antigen (PSA) was 5,000 ng/ml or higher. Transrectal needle biopsy of the prostate revealed moderately differentiated adenocarcinoma. Computed tomography (CT) and bone scan showed para-aorta lymph node metastasis and bone metastasis. Hormone therapy was performed. Case 2; a 37-year-old man admitted with the chief complaint of pollakisuria and sense of residual urine. Serum level of prostate-specific antigen (PSA) was 24 ng/ml. Magnetic resonance imaging (MRI) showed that the prostate tumor invaded the bladder wall. Transrectal needle biopsy revealed poorly differentiated adenocarcinoma. Hormone therapy and radiation therapy were performed. Twenty-one cases reported in Japan in addition to the present cases are reviewed.

[Mucinous adenocarcinoma of the prostate: a case report and review of 32 cases on immunohistochemical study of both PSA and CEA].

Mucinous adenocarcinoma of the prostate is extremely rare and its biological behavior is not well known. We report a case of mucinous adenocarcinoma of the prostate which stained positively for prostate specific antigen (PSA) and negatively for carcinoembryonic antigen (CEA) on immunohistochemical study. Our case contained conventional adenocarcinomas and no signet-ring cells. Thirty two cases of mucinous adenocarcinoma of the prostate which performed on immunohistochemical study of both PSA and CEA, including our case, were reviewed. 17 of the 23 cases of immunoreactive to PSA contained conventional adenocarcinomas, and 3 of the 10 cases of immunoreactive to CEA contained them, respectively. The 6 cases of immunoreactive to CEA only contained signet-ring cells. It indicated that there seemed to be the two types of mucinous adenocarcinoma of the prostate, the one which stained positively for PSA was the subtype of conventional adenocarcinomas, and the other which stained positively for CEA and negatively for PSA was derived from the intestinal metaplasia with atypia of the prostatic urethra.

[Retrospective analysis of chief complaints of patients with urogenital malignancies over the past decade at the Jikei University Hospital].

We analyzed the chief complaints of patients with four major urogenital malignancies (renal cancer, renal pelvis and ureter cancer, bladder cancer and prostatic cancer) over the past decade (1990-1999) at the Jikei University Hospital. Over the last 10 years, a high percentage of renal cancers were detected incidentally. By contrast, prostatic cancers were more likely (10.5%) than other cancers to be detected on the basis of symptoms of metastasis. However, since 1995 more prostatic cancers are being detected with prostatic-specific antigen screening at the health checkups. Gross hematuria is the chief complaint of most patients with uroepithelial cancers (cancers of the renal pelvis, ureter and bladder cancer). Additionally, renal pelvis and ureter cancers were diagnosed with screening in a few patients in the past five years.

Need for cytochrome bc1 complex for dissimilatory nitrite reduction of Pseudomonas aeruginosa.

Pseudomonas aeruginosa strains deficient in the genes for cytochrome c1, a subunit of the cytochrome bc1 complex, or the tetraheme membrane protein NapC, which is similar to NirT of Pseudomonas stutzeri, were constructed and their growth was investigated. The cytochrome c1 mutant could not grow under anaerobic conditions with nitrite as an electron acceptor and did not reduce nitrite in spite of its producing active nitrite reductase. NirM (cytochrome c551) and azurin, which are the direct electron donors for nitrite reductase, were reduced by succinate in the presence of the membrane fraction from the wild-type strain as a mediator but not in the presence of that from the cytochrome c1 mutant. These results indicated that cytochrome bc1 complex was necessary for electron transfer from the membrane quinone pool to nitrite reductase. The NapC mutant grew anaerobically at the expense of nitrite, indicating that NapC was not necessary for nitrite reduction.

[Fatal septic shock caused by transrectal needle biopsy of the prostate; a case report].

A 46-year-old man refer to us because of hemospermia. The prostatic gland was normal in size and consistency at rectal examination. Serum prostate specific antigen was 7.04 ng/ml. Magnetic resonance imaging showed an area of low signal intensity on T2-weighted images in the left peripheral gland, possibly indicative of carcinoma. Transrectal prostate biopsy was performed after intravenous administration of piperacillin. He developed chills and fever (39 degrees C) the next morning following biopsy. He was taken unconscious into the hospital where a diagnosis of septic shock caused by Escherichia coli was made. Five days later, he died. His general condition deteriorated notwithstanding intensive treatment. Postmortem blood cultures were positive for a piperacillin resistant Escherichia coli. Histological examination of the biopsies showed a benign prostatic hyperplasia. Autopsy showed diffuse tissue damage in the heart, lung, liver and kidneys. The prostate had numerous microabscesses. Currently, transrectal prostate biopsy is considered a generally reliable procedure to detect adenocarcinoma of the prostate. Our case seems to the sixth case report of fatal complications.

The role of renal scintigraphy in renal arteriovenous malformation treated by transcatheter arterial embolization.

A 41-year-old woman was hospitalized because of gross hematuria. She was found to have a renal arteriovenous malformation and was treated with transcatheter arterial embolization (TAE). Renal scintigraphy with Tc-99m mercaptoacetyltriglycine was performed before and after TAE. Renal scintigraphy was useful for evaluating renal function in this patient.