The effect of mutation on an aggregation-prone protein: An in vivo, in vitro, and in silico analysis.

Aggregation of initially stably structured proteins is involved in more than 20 human amyloid diseases. Despite intense research, however, how this class of proteins assembles into amyloid fibrils remains poorly understood, principally because of the complex effects of amino acid substitutions on protein stability, solubility, and aggregation propensity. We address this question using ß2-microglobulin (ß2m) as a model system, focusing on D76N-ß2m that is involved in hereditary amyloidosis. This amino acid substitution causes the aggregation-resilient wild-type protein to become highly aggregation prone in vitro, although the mechanism by which this occurs remained elusive. Here, we identify the residues key to protecting ß2m from aggregation by coupling aggregation with antibiotic resistance in E. coli using a tripartite ß-lactamase assay (TPBLA). By performing saturation mutagenesis at three different sites (D53X-, D76X-, and D98X-ß2m) we show that residue 76 has a unique ability to drive ß2m aggregation in vivo and in vitro. Using a randomly mutated D76N-ß2m variant library, we show that all of the mutations found to improve protein behavior involve residues in a single aggregation-prone region (APR) (residues 60 to 66). Surprisingly, no correlation was found between protein stability and protein aggregation rate or yield, with several mutations in the APR decreasing aggregation without affecting stability. Together, the results demonstrate the power of the TPBLA to develop proteins that are resilient to aggregation and suggest a model for D76N-ß2m aggregation involving the formation of long-range couplings between the APR and Asn76 in a nonnative state.

The Chennai Declaration: a roadmap to tackle the challenge of antimicrobial resistance.

"A Roadmap to Tackle the Challenge of Antimicrobial Resistance - A Joint meeting of Medical Societies in India" was organized as a pre-conference symposium of the 2 nd annual conference of the Clinical Infectious Disease Society (CIDSCON 2012) at Chennai on 24 th August. This was the first ever meeting of medical societies in India on issue of tackling resistance, with a plan to formulate a road map to tackle the global challenge of antimicrobial resistance from the Indian perspective. We had representatives from most medical societies in India, eminent policy makers from both central and state governments, representatives of World Health Organization, National Accreditation Board of Hospitals, Medical Council of India, Drug Controller General of India, and Indian Council of Medical Research along with well-known dignitaries in the Indian medical field. The meeting was attended by a large gathering of health care professionals. The meeting consisted of plenary and interactive discussion sessions designed to seek experience and views from a large range of health care professionals and included six international experts who shared action plans in their respective regions. The intention was to gain a broad consensus and range of opinions to guide formation of the road map. The ethos of the meeting was very much not to look back but rather to look forward and make joint efforts to tackle the menace of antibiotic resistance. The Chennai Declaration will be submitted to all stake holders.

Primary chondrosarcoma of the breast: a case report.

Mammary sarcomas are uncommon tumors. Of these pure chondrosarcoma without any other area of epithelial or mesenchymal differentiation feature as even rarer. This excludes tumors like malignant cystosarcoma phyllodes and metaplastic carcinoma where malignant cartilaginous areas may be present. Further primary chondrosarcoma arises from breast stroma and not from underlying bone or cartilage. Only five cases of pure and primary chondrosarcoma have been reported so far. The sixth case is reported here.

Models for environmentally induced eating disorders: dietary hyperphagia and anorexia nervosa.

The two protocols in this unit provide suggestions for constructing models of eating disorders that are at the opposite ends of the spectrum: dietary hyperphagia and anorexia nervosa. The greatest degree of dietary hyperphagia is induced by giving rats or mice access to a daily choice of highly palatable foods (e.g., chocolate or bread) in addition to their regular chow. Like humans, rats overeat and actually develop physiological requirements for these foods. This model can be used to test the effects of putative anorectic agents on both acute and chronic administration regimens. The second protocol describes a model of compulsive behavior that results in profound weight loss, which is produced by moderate food deprivation along with continuous access to exercise wheels.

Adenocarcinoma renal: factores pronósticos: Experiencia de 10 años del club urológico del sur de la Provincia de Buenos Aires (C.U.S.P.B.A) / Renal adenocarcinoma: prognostic factors: 10 years experience of the Provincia Buenos Aires urologic Club

Objetivo: Establecer factors pronósticos en el adenocarcinoma renal.Material y método: Se realizó un estudio retrospectivo de todos los casos de adenocarcinoma renal operados por los miembros del Club urológico del sur de la provincia de Buenos Aires que participaron en este estudio, entre los años 1986 y 1996. Resultados: Se revisaron 231 casos de carcinoma renal operados en ese período. La edad fue de 62+= 11,63 por ciento fueron hombres. La sobrevida a 5 años fue : 100 por ciento estadio I, 77 por ciento estadio II, 51 por ciento estadio III y, 0 por ciento estadio IV; 32 por ciento fueron incidentales. Los pacientes < de 60 años y los tumores > de 5 cm presentaronmenor sobrevida. Conclusiones: Los tumores de bajo estadio (I y II), < de 5 cm, y en pascientes mayores de 60 años tienen mejor pronóstico en cuanto a la sobrevida global

Interstitial laser hyperthermia, a new method in the management of fibroadenoma of the breast: A pilot study.

BACKGROUND AND OBJECTIVE: This study attempts to evaluate the effect of interstitial laser hyperthermia in breast fibroadenomas as an outpatient procedure. STUDY DESIGN/MATERIALS AND METHODS: In an uncontrolled prospective study, 27 patients younger than 35 years were subjected to laser phototherapy of their breast fibroadenomas. Under real-time ultrasound monitoring, Nd:YAG laser (1,064 nm wavelength) was used at 2 W for 300 sec (600 J) in a continuous wave mode to produce interstitial hyperthermia. Follow-ups were done at 2, 4, and 8 weeks. Subsequently, excision biopsy of residual lumps was performed. RESULTS: There was significant decrease in clinical and sonographic sizes (P < 0.001). Follow-up ultrasound showed a progressive change of hyperechoic texture, from a heterogeneous to a nearly homogeneous one. There were minimal scars (2-3 mm) and no keloid or abscess formation. CONCLUSION: Interstitial laser hyperthermia is a safe, precise, and minimally invasive outpatient procedure for in situ destruction of breast fibroadenomas.

Insuficiencia renal en pacientes nefrectomizados por cancer. Experiencia de diez años del Club Urológico del Sur de la Provincia de Buenos Aires / Renal failure in patients who undergone nephrectomy because of cancer. Experience of CUSPBA

Objetivo: Establecer la incidencia de insuficiencia renal posoperatoria en pacientes nefrectomizados por adenocarcinoma renal. Material y método: Se analizaron retrospectivamente todas las nefrectomías por cancer renal realizadas por los participantes en el estudio entre los años 1986 y 1996. Se evaluó la presencia de insuficiencia renal pre y posoperatoria tomando valores de creatinina de 1,5 mg/dl como límite superior. Se evaluó si hubiera sido posible realizar cirugía conservadora en cada caso. Resultados: se revisaron 231 casos de nefrectomía por cáncer. El 7,8 por ciento presentó insuficiencia reanl posoperatoria (3,9 por ciento transitoria y 3,9 por ciento definitiva) con creatininas>1,5 mg/dl. Sólo el 0,43 por ciento requirió hemodiálisis crónica. La falla renal se presentó a los 18,7 meses posoperatorios. Dentro del grupo donde se hubiera podido efectuar cirugía conservadora, el 2,1 por ciento presentó insuficiencia renal definitiva. Conclusiones: La incidencia de falla renal posnefrectomía es del 7,8 por ciento(3,9 definitiva y 3,9 por ciento transitoria). Sólo el 0,43 por ciento requiere hemodiálisis crónica

Cytotoxic effects of 125I-labeled PBZr ligand PK 11195 in prostatic tumor cells: therapeutic implications.

The effect of [125I]PK 11195 was examined in human prostatic tumor cells (DU 145) in culture and compared with Na[125I] and non-radioactive PK 11195. [125I]PK 11195 was clearly cytocidal. The data for dose-related cell survival with [125I]PK 11195 showed a linear relationship. Na[125I] or non-labeled PK 11195 at similar concentrations did not lead to any cell killing. The uptake of [125I]PK 11195 and [3H]PK 11195 in cells was very similar. Fragmentation of DNA measured by agarose gel electrophoresis showed that exposure of DU 145 cells to [1251]PK 11195 for 1, 4 or 24 h caused no fragmentation. These results indicate that nuclear DNA is not the prime binding site for [125I]PK 11195, which is consistent with the presence of specific peripheral benzodiazepine receptors (PBZr) in the mitochondria. The cell killing effect of [125I]PK 11195 suggests the use of PBZr ligand for radiotherapy.

Allelic forms of the knob associated histidine-rich protein gene of Plasmodium falciparum.

The knob associated histidine-rich protein (KAHRP) gene was cloned and sequenced from two Indian isolates of Plasmodium falciparum, Pf3-92 and Pf29-92. These isolates showed major sequence differences in the C-terminal repeat domain of KAHRP. However, the biologically important domains such as spectrin-actin binding region remained highly conserved. The PCR amplification of a variable C-terminal repeat domain from the clinical isolates of P. falciparum, from Rajasthan epidemic, showed the presence of multiple alleles of KAHRP gene. The presence of multiple alleles indicates the existence of several P. falciparum strains in India. This should be taken into account for future malaria control strategies such as molecular therapy and vaccines.

Hepatic metastases: interstitial laser photocoagulation with real-time US monitoring and dynamic CT evaluation of treatment.

Fifty-five liver metastases in 21 patients were treated with interstitial laser photocoagulation (ILP). Tumors were irradiated with a neodymium yttrium aluminum garnet laser via optical fibers passed through 19-gauge needles inserted under ultrasound (US) guidance. Heating of the tumor was evident at real-time US as an expanding and coalescing echogenic zone around the needle tips. After ILP, dynamic computed tomography (CT) showed laser-induced necrosis as a new area of nonenhancement. Necrosis of tumor volume was more than 50% in 82% (45 of 55) of the tumors, and 100% necrosis was achieved in 38% (21 of 55). Metastases smaller than 4 cm in diameter were treated more effectively and required fewer treatment sessions than did those larger than 4 cm. Complications were minor and included severe pain in four cases, persistent pain for up to 10 days in 11 cases, and asymptomatic subcapsular hematoma (four cases) and pleural effusion (six cases) seen with CT. ILP is safe and effective for liver tumor destruction, and US and CT are useful in different aspects of treatment monitoring.