Correlation of expression of the actin filament-bundling protein espin with stereociliary bundle formation in the developing inner ear

The vertebrate hair cell is named for its stereociliary bundle or hair bundle that protrudes from the cell's apical surface. Hair bundles mediate mechanosensitivity, and their highly organized structure plays a critical role in mechanoelectrical transduction and amplification. The prototypical hair bundle is composed of individual stereocilia, 50-300 in number, depending on the animal species and on the type of hair cell. The assembly of stereocilia, in particular, the formation during development of individual rows of stereocilia with descending length, has been analyzed in great morphological detail. Electron microscopic studies have demonstrated that stereocilia are filled with actin filaments that are rigidly cross-linked. The growth of individual rows of stereocilia is associated with the addition of actin filaments and with progressively increasing numbers of cross-bridges between actin filaments. Recently, a mutation in the actin filament-bundling protein espin has been shown to underlie hair bundle degeneration in the deaf jerker mouse, subsequently leading to deafness. Our study was undertaken to investigate the appearance and developmental expression of espin in chicken inner ear sensory epithelia. We found that the onset of espin expression correlates with the initiation and growth of stereocilia bundles in vestibular and cochlear hair cells. Intense espin immunolabeling of stereocilia was colocalized with actin filament staining in all types of hair cells at all developmental stages and in adult animals. Our analysis of espin as a molecular marker for actin filament cross-links in stereocilia is in full accordance with previous morphological studies and implicates espin as an important structural component of hair bundles from initiation of bundle assembly to mature chicken hair cells.     

Antisense therapy specific to mutated K-ras gene in hamster pancreatic cancer model. Can it inhibit the growth of 5-FU and MMC-resistant metastatic and remetastatic cell lines?

K-ras point mutation at codon 12 has a relationship greater than 90% with pancreatic cancer. Cancer therapy should also include the treatment of metastatic disease because it is known that the properties of metastatic cells may vary considerably from those of the primary tumor. AIM: To clarify if the same drugs, which can inhibit the tumor growth in the parental cell line, can inhibit the pancreatic metastatic and remetastatic cell lines at the same concentrations and to compare the inhibition with antisense oligonucleotides mismatched to K-ras gene, in Syrian golden hamsters. MATERIALS AND METHODS: HaP-T1, a BHP-induced hamster pancreatic cancer cell line, MS-PaS-1 (a metastatic cell line established from "return trip" metastases from the liver to the pancreas) and MS-PaS-2 named as a "remetastatic cell line", i.e., metastases from MS-PaS-1 were used. MTT and MTT-agarose assays were performed, using 5-Fluorouracil (5-FU), Mitomycin C (MMC) and antisense oligonucleotide specific to K-ras oncogene. RESULTS: The inhibitory concentration (IC50) of 5-FU, which inhibited HaP-T1, had to be increased by 50-fold to inhibit MS-PaS-1 and 100-fold to inhibit MS-PaS-2. MMC had to be increased by 10-fold to inhibit MS-PaS-1 and 50-fold to inhibit MS-PaS-2. However, IC50 was the same when antisense oligonucleotide was tried in these 3 cell lines. CONCLUSION: Antisense oligonucleotide-targeted K-ras gene may be a good choice for therapy because it could inhibit the growth in metastatic and remetastatic cells as well as in primary tumor cells.     

Assessment of the efficacy of joint lavage versus joint lavage plus corticoids in patients with osteoarthritis of the knee

PURPOSE: Joint lavage (JL), involves the passage of cold sterile 0.9% saline through the knee joint in order to have the fluid reach the inside of the joint capsule. This technique was evaluated as a local treatment for osteoarthritis (OA) of the knee alone (JL) and in combination with intra-articular infiltration with glucocorticoids (JLC). PATIENTS AND METHODS: An overall 299 knees belonging to 205 patients (22% males, 78% females) with a mean age of 67 +/- 8 years and osteoarthritis of the knee of radiological grade II or III on the Kellgren scale were randomised in the ratio of 1:4 into two therapeutic groups, namely: JL (n = 62) and JLC (n = 237). All patients received joint lavage on day 0; in those of the JLC group, joint lavage was followed by infiltration of 40 mg of triamcinolone acetonide. The efficacy of both treatments was assessed by recording the corresponding values for the following variables: pain strength as measured by a visual analogy scale (VAS), effusion, crepitation, restricted motion, of osteoarthritis of the knee. spontaneous pain, pain on pressure, pain on passive motion and pain on active motion; all of these were recorded at the onset of the study, and after 1 and 3 months. RESULTS: There were no significant differences in the values of the variables at the different followup times. Also, pain severity was similar in both treatment groups. Thus, VAS for pain was 7.3 +/- 0.3 for the JL group and 7.1 +/- 0.2 for the JLC group at the onset, and decreased to 3.0 +/- 0.3 in the former and 2.8 +/- 0.2 in the latter after 1 month; the decrease was statistically significant in both cases. After 3 months, the JL and JLC groups had a VAS of 3.5 +/- 0.3 and 3.8 +/- 0.2, respectively. CONCLUSIONS: The results of this work suggest the absence of significant differences between the two treatments, such that both joint lavage alone and with infiltration with corticoids can be concluded as similarly effective for the symptomatic management     

Embryotoxicity in vitro with extract of Indigofera suffruticosa leaves

Aqueous extract of leaves of Indigofera suffruticosa (AELIs) were studied for adverse effects in preimplantation mouse embryos. Two-cell mouse embryos were cultured for 94 h in human tubal fluid medium (HTF), and AELIs at a concentration of 5 or 10 mg/ml. On Day 4 of culture, morulae and blastocysts were collected for morphological analysis of blastomeres. We found that embryos exposed to the higher concentration of AELIs (10 mg/ml) did not develop and all embryos persisted at the two-cell stage. Those embryos exposed to the lower concentration (5 mg/ml) showed development until morula, blastocyst and hatched blastocyst stages that were similar to the controls. These results suggest that use of AELIs may be hazardous to humans who make use of it in folk medicine.     

Comparison between C57Bl/6 and C57Bl/10 mycobacterial mouse pleurisy with respect to cellular migration and nitric oxide production

Mycobacterium bovis-BCG (BCG) and Mycobacterium leprae (ML) have opposite inflammatory properties. Mycobacteria-induced pleurisy in C57Bl/6 and C57Bl/10 mice was evaluated to establish if their innate responses could be comparable, verifying cellular migration and nitrite production. Kinetic responses after ML or BCG intrathoracic injection were compared in those mice, sharing the H-2^b MHC haplotype. BCG led to acute eosinophilia and late neutrophilia in both mice. In C57Bl/6 late pleurisy, monocytes and neutrophil recruitment was dose- and iNOS-dependent, inhibited by methotrexate but not by indomethacin. Pleural macrophages released nitrites ex vivo after 7 days of BCG stimulus, without “priming” and blocked by the nitrite inhibitor L-N5-(1-iminoethyl)-ornithine (L-NIO). ML did not induce cellular migration or nitrite production, independent of the mouse strain, timing, or number of bacilli. Although these mycobacteria have high homology, there was no effect of ML on BCG-evoked secondary cellular recruitment. Both C57Black mice trigger similar onset of inflammatory responses to these mycobacteria, so far can alternatively be used in experimental studies.     

Environmental temperature, cholera, and acute diarrhoea in adults in Lima, Peru

The effects of environmental temperature, presence and severity of El Ni?o, presence of cholera in the community, and interactions among these variables on the number of adult diarrhoeal patients attending the Hospital Nacional Cayetano Heredia in Lima, Peru, during 1991-1998, were evaluated. During 1991-1996, an increased number of visits to the hospital due to acute diarrhoea in the warmer months was observed. This periodic pattern was altered in 1997, when rising of the environmental temperature was observed in Lima secondarily associated with a strong El Ni?o event. A multivariate model was built in which environmental temperature and interaction between environmental temperature and presence of cholera predicted the number of adult patients with acute diarrhoea attending the Hospital Nacional Cayetano Heredia. Monitoring of environmental temperature and presence of cholera may be used as a warning system to predict epidemics of diarrhoea in adults, which may have a tremendous impact on healthcare strategies and management of health services in general.     

In vitro investigation of voriconazole susceptibility for keratitis and endophthalmitis fungal pathogens

PURPOSE: To update the spectrum of ocular fungal isolates and investigate the in vitro efficacy of voriconazole and other antifungals. DESIGN: Experimental study. METHODS: Microbiology database was scanned and fungal isolates associated with keratitis (419) and endophthalmitis (122) were analyzed for classification and isolate frequency. The Sensititre YeastOne microdilution antifungal susceptibility test was used to evaluate susceptibility (MICs) of 34 common fungal pathogens against amphotericin B, fluconazole, ketoconazole, 5-flucytosine, itraconazole, and voriconazole. Ten of the test isolates were sent to a reference laboratory to validate the Sensititre results. RESULTS: Fusarium species remains the most frequent corneal fungal pathogen (60.1%). Colletotrichum species (4.1%) has emerged as the fifth most common mold in keratitis. Top yeast isolates from cornea included Candida albicans (52.3%) and Candida parapsilosis (37.3%). Half of the intraocular pathogens were Candida species. Paecilomyces (2.9%) and Philophora (1.9) were unusual pathogens. In vitro susceptibility profiles were voriconazole (100%), ketoconazole (82.4%), amphotericin (76.5%), itraconazole (67%), fluconazole (60%), and 5-FC (60%). Voriconazole MIC(90) were lowest for Candida species (0.016 microg/ml) and highest for Fusarium species (2 microg/ml). Reference laboratory MICs correlated 100% for yeast isolates (0.016 microg/ml) but were fourfold higher for Fusarium species (8 microg/ml). MIC(90) for Aspergillus species was 0.5 microg/ml. CONCLUSIONS: Candida, Fusarium, and Aspergillus species remain frequent fungal pathogens. Voriconazole may have a role in the therapeutic management of Candida and Aspergillus ocular infections. Clinical efficacy must determine the role for other fungal pathogens. Human use and animal models will determine its use in the clinical setting.     

A modified technique to stain the internal limiting membrane with indocyanine green

PURPOSE: Peeling of the internal limiting membrane (ILM) has improved the outcomes in vitreoretinal surgery of macular diseases. Indocyanine green (ICG) is used to stain and improve the visualization of the ILM. This paper aims to describe a modified approach to stain and peel the ILM avoiding potential adverse side effects. METHODS: After a core vitrectomy, 0.05 ml of ICG in a concentration of 5 mg/ml is quickly injected under water around the macular region. The tip of the syringe is positioned about 5 mm from the macular tissue, in a way that about a 3-mm diameter area around the fovea is stained by the ICG. Additional ICG is immediately removed by aspiration. Peeling of the ILM is accomplished with a forceps. RESULTS: Neither residual ICG in the vitreous cavity nor any clinical signs of phototoxicity like retinal edema were detected in any patients operated by this technique. CONCLUSIONS: This modified technique to stain the ILM limits the amount and concentration of ICG. The locally limited contact of ICG with the retinal surface seems to be a safe procedure to stain the ILM.