Differential Effects of Trp53 Alterations in Murine Colorectal Cancer.

BACKGROUND: Colorectal cancer (CRC) development is a multi-step process resulting in the accumulation of genetic alterations. Despite its high incidence, there are currently no mouse models that accurately recapitulate this process and mimic sporadic CRC. We aimed to develop and characterize a genetically engineered mouse model (GEMM) of Apc/Kras/Trp53 mutant CRC, the most frequent genetic subtype of CRC. METHODS: Tumors were induced in mice with conditional mutations or knockouts in Apc, Kras, and Trp53 by a segmental adeno-cre viral infection, monitored via colonoscopy and characterized on multiple levels via immunohistochemistry and next-generation sequencing. RESULTS: The model accurately recapitulates human colorectal carcinogenesis clinically, histologically and genetically. The Trp53 R172H hotspot mutation leads to significantly increased metastatic capacity. The effects of Trp53 alterations, as well as the response to treatment of this model, are similar to human CRC. Exome sequencing revealed spontaneous protein-modifying alterations in multiple CRC-related genes and oncogenic pathways, resulting in a genetic landscape resembling human CRC. CONCLUSIONS: This model realistically mimics human CRC in many aspects, allows new insights into the role of TP53 in CRC, enables highly predictive preclinical studies and demonstrates the value of GEMMs in current translational cancer research and drug development.

Onychomycosis due to Achaetomium strumarium.

Onychomycosis is the most common infection of the toe-nails or finger-nails and it may be caused by a large variety of fungal species. Achaetomium species which belong to the phylum Ascomycota (Family Chaetomiaceae), are usually soil saprophytes or endophytic fungi which have been rarely reported as human or animal pathogens. Here, we report a case of onychomycosis caused by Achaetomium strumarium in a healthy person who showed involvement of all fingers of both hands with yellowish brown discoloration. The causative agent isolated was identified as Achaetomium species by morphology, colony morphometry and growth at high temperature and as A. strumarium from DNA sequence of ITS region. Onychomycosis from this case responded satisfactorily with per os (P. O.; oral) and topical application of Terbinafine.

A Genetically Engineered Mouse Model of Sporadic Colorectal Cancer.

Despite the advantages of easy applicability and cost-effectiveness, colorectal cancer mouse models based on tumor cell injection have severe limitations and do not accurately simulate tumor biology and tumor cell dissemination. Genetically engineered mouse models have been introduced to overcome these limitations; however, such models are technically demanding, especially in large organs such as the colon in which only a single tumor is desired. As a result, an immunocompetent, genetically engineered mouse model of colorectal cancer was developed which develops highly uniform tumors and can be used for tumor biology studies as well as therapeutic trials. Tumor development is initiated by surgical, segmental infection of the distal colon with adeno-cre virus in compound conditionally mutant mice. The tumors can be easily detected and monitored via colonoscopy. We here describe the surgical technique of segmental adeno-cre infection of the colon, the surveillance of the tumor via high-resolution colonoscopy and present the resulting colorectal tumors.

Keratitis by a rare pathogen Colletotrichum gloeosporioides: A case report.

Colletotrichum species have been reported infrequently as the cause of keratitis or subcutaneous lesions. The patient we describe developed keratitis after ocular trauma. The sample from the corneal scrapings grew Colletotrichum gloeosporioides as identified from morphological characters and DNA sequence of the 'Internal Transcribed Spacer' (ITS) region. The patient underwent topical application of amphotericin-B followed by itraconazole and natamycin treatment. Simultaneous oral voriconazole regimen leads to complete regression of corneal ulcer. This report highlights the fact that early and accurate identification and therapy can resolve keratitis caused by rare pathogen C. gloeosporioides.

Epidemiological evaluation of acute gastroenteritis and therapeutic approaches in Middle East Countries.

OBJECTIVE: Gastroenteritis represents with respiratory tract infection the most common infectious disease syndrome of humans in developing countries. Gut microbiota regional variation and dysbiosis play a crucial role in triggering and worsening this devastating GI disease. MATERIALS AND METHODS: With this manuscript, we want to explore and emphasize the critical aspect of acute gastroenteritis in Middle-East Countries and its correlation with the clinical aspect of gut microbiota modification and intestinal homeostasis. RESULTS: Approximately 1 of 50 children born each year in developed nations is hospitalized for acute gastroenteritis sometime during childhood. The highest rate of illness occurs in children between 3 and 24 months of age. The common causes of diarrhea are infections with viruses and bacteria, diarrhea due to a systemic infection other than gastrointestinal, diarrhea associated with antibiotic administration, and feeding related diarrhea. The single most common diarrheal disorder observed in the Emergency Department and general practice is viral gastroenteritis. In particular, Rotavirus is the cause of more than 2 million hospitalizations and over half a million deaths from acute GE in infants and young children. This burden produces also direct and indirect economic costs. The use of probiotics to counterbalance commensal dysbiosis is emerging as a standard medical practice in these countries. CONCLUSIONS: In this scenario, one of the most interesting aspects is that probiotics and gut microbiota modulation could deeply improve the prevention and treatment of this devastating GI pathology. At the same time, vaccination might represent a cost-effective strategy to reduce the health and economic burden of some pathogens, such as rotavirus.

Circulating tumor cells exhibit stem cell characteristics in an orthotopic mouse model of colorectal cancer.

The prognosis of colorectal cancer (CRC) is closely linked to the occurrence of distant metastases, which putatively develop from circulating tumor cells (CTCs) shed into circulation by the tumor. As far more CTCs are shed than eventually metastases develop, only a small subfraction of CTCs harbor full tumorigenic potential. The aim of this study was to further characterize CRC-derived CTCs to eventually identify the clinically relevant subfraction of CTCs.We established an orthotopic mouse model of CRC which reliably develops metastases and CTCs. We were able to culture the resulting CTCs in vitro, and demonstrated their tumor-forming capacity when re-injected into mice. The CTCs were then subjected to qPCR expression profiling, revealing downregulation of epithelial and proliferation markers. Genes associated with cell-cell adhesion (claudin-7, CD166) were significantly downregulated, indicating a more metastatic phenotype of CTCs compared to bulk tumor cells derived from hepatic metastases. The stem cell markers DLG7 and BMI1 were significantly upregulated in CTC, indicating a stem cell-like phenotype and increased capacity of tumor formation and self-renewal. In concert with their in vitro and in vivo tumorigenicity, these findings indicate stem cell properties of mouse-derived CTCs.In conclusion, we developed an orthotopic mouse model of CRC recapitulating the process of CRC dissemination. CTCs derived from this model exhibit stem-cell like characteristics and are able to form colonies in vitro and tumors in vivo. Our results provide new insight into the biology of CRC-derived CTCs and may provide new therapeutic targets in the metastatic cascade of CRC.

Idiopathic pulmonary hemosiderosis: alveoli are an answer to anemia.

Idiopathic pulmonary hemosiderosis (IPH) is a rare disorder (triad of iron-deficiency anemia, hemoptysis, and alveolar infiltrates). A 3-year-old male presented with mild fever, breathlessness, dry cough, and bluish nail discoloration for 8 days. He had required five blood transfusions in the past 1 year (last transfusion was given 4 months ago). He had a respiratory rate of 58/min with respiratory distress, cyanosis, and grade III clubbing. Respiratory system examination was normal. Several previous reports of hemoglobin were as low as 3.6 g/dl with hypochromic and microcytic anemia. There were transient increases in the hemoglobin and normalization of red cell morphology with blood transfusions. Serum iron, G6PD enzyme assay, hemoglobin electrophoresis, the sickling test, Coomb's test, stool and urine analysis, and a Meckel's scan were normal. HIV antibody and dsDNA were negative. The chest radiograph revealed symmetrical patchy infiltrates sparing lung apices (confirmed on high-resolution computed tomography). Lung biopsy diagnosed pulmonary hemosiderosis (interstitial lung disease with hemosiderin-laden macrophages scattered in the alveoli and areas of fibrosis in the alveolar septa). The patient showed marked clinical improvement in 10 days of therapy with prednisolone. IPH should be listed in the differential diagnosis of a child presenting with unexplained hypochromic, microcytic anemia and respiratory symptoms.

Seronegative necrolytic acral erythema: a distinct clinical subset?

A patient was referred to us with asymptomatic, erythematous, nonitchy, scaly lesions present bilaterally on the dorsa of his feet and toes since the last 2 months. Both the legs had pitting edema as well. There were hyperkeratosis, focal parakeratosis, acanthosis and scattered spongiosis in the epidermis, and proliferation of capillaries with perivascular infiltration of lymphomononuclear cells in the dermis. There was no serological evidence of hepatitis C virus. Laboratory investigations revealed hypoalbuminemia and low-normal serum zinc. On clinicopathological correlation, we made a diagnosis of necrolytic acral erythema (NAE). The lesions responded dramatically to oral zinc sulfate and topical clobetasol propionate within 3 weeks with disappearance of edema and scaling and only a minimal residual erythema. This is the first reported case of NAE from Eastern India. NAE with negative serology for hepatitis C may be viewed as a distinct subset of the condition that had been originally described.

Neurotoxoplasmosis mimicking intracranial tuberculoma.

Neurotoxoplasmosis is a major cause of morbidity and mortality, especially in immunocompromised patients. Definitive diagnosis is invasive and difficult thereby requiring a therapeutic trial. We herein report a case of an 11-year-old boy who presented with a tuberculoma on computerized tomography (CT) brain and did not show any clinical improvement on anti-tuberculous drugs. Subsequently, reviewing the CT scans and with supportive serology, a diagnosis of neurotoxoplasmosis was considered. A trial of antitoxoplasmosis therapy was given to which the child fully responded in two weeks. Three weeks later, his magnetic resonance imaging showed complete resolution of the lesion. This report provides an insight into the significance of therapeutic trial in neurotoxoplasmosis.

Validation of microarray-based resequencing of 93 worldwide mitochondrial genomes.

The human mitochondrial genome consists of a multicopy, circular dsDNA molecule of 16,569 base pairs. It encodes for 13 proteins, two ribosomal genes, and 22 tRNAs that are essential in the generation of cellular ATP by oxidative phosphorylation in eukaryotic cells. Germline mutations in mitochondrial DNA (mtDNA) are an important cause of maternally inherited diseases, while somatic mtDNA mutations may play important roles in aging and cancer. mtDNA polymorphisms are also widely used in population and forensic genetics. Therefore, methods that allow the rapid, inexpensive and accurate sequencing of mtDNA are of great interest. One such method is the Affymetrix GeneChip Human Mitochondrial Resequencing Array 2.0 (MitoChip v.2.0) (Santa Clara, CA). A direct comparison of 93 worldwide mitochondrial genomes sequenced by both the MitoChip and dideoxy terminator sequencing revealed an average call rate of 99.48% and an accuracy of > or =99.98% for the MitoChip. The good performance was achieved by using in-house software for the automated analysis of additional probes on the array that cover the most common haplotypes in the hypervariable regions (HVR). Failure to call a base was associated mostly with the presence of either a run of > or =4 C bases or a sequence variant within 12 bases up- or downstream of that base. A major drawback of the MitoChip is its inability to detect insertions/deletions and its low sensitivity and specificity in the detection of heteroplasmy. However, the vast majority of haplogroup defining polymorphism in the mtDNA phylogeny could be called unambiguously and more rapidly than with conventional sequencing.

Synthesis of bis-armed amino acid derivatives via the alkylation of ethyl isocyanoacetate and the Suzuki-Miyaura cross-coupling reaction.

Two synthetic routes to bis-armed-alpha-amino acid derivatives are described. The first route involves alkylation of dibromo derivatives with ethyl isocyanoacetate under phase-transfer catalysis (PTC) conditions. The second route uses a palladium-mediated Suzuki-Miyaura cross-coupling reaction between a DL-4-boronophenylalanine derivative and aromatic diiodo (or dibromo) compounds.

Hepatitis B e Antigen Negative Chronic Hepatitis in Indian Patients : A Reality.

BACKGROUND: Hepatitis B e antigen negative chronic hepatitis (e(-) CHB) with detectable levels of hepatitis B virus DNA (HBV DNA) in serum has been reported in cases from Asia. This study was undertaken to find out prevalence e(-)CHB and to correlate its presence with the clinical status and severity of the illness in cases of chronic liver disease in India. METHODS: All patients of infective hepatitis, who were hepatitis B surface antigen (HBsAg) positive by enzyme-linked immunosorbent assay (ELISA), were evaluated with liver function tests and HBeAg and antiHBe antibody studies. Polymerase chain reaction (PCR) test was carried out to detect HBV DNA qualitatively. RESULT: Out of 2064 samples tested by ELISA, 429 (20.78 %) were HBsAg positive. HBV DNA (qualitative) was performed on all 429 patients and 74 (17.2%) were HBV DNA positive. Of these only 42 (56.75 %) tested positive for HBeAg. Overall, 8.3 % of HBeAg negative patients (32/384) were viraemic with evidence of chronic liver disease/clinical cirrhosis and alteration of transaminase levels, while three cases (0.84 %) HBeAg positive cases did not show presence of HBV DNA. CONCLUSION: This study shows e(-)CHB prevalence rate of 8.3% in patients with HBV infection in India. Since HBeAg negative patients had detectable levels of HBV DNA as seen in HBeAg positive patients, benefit of antiviral therapy should be given to them. Population studies on e(-) CHB cases are needed to determine its true prevalence, natural course and response to therapy.

Pseudomonas sepsis with Noma: an association?

We report here a 2.5-year-old male child with community-acquired Pseudomonal sepsis showing the characteristic lesions of ecthyma gangrenosum. The child had development of gangrenous changes of the nose and face - the 'cancrum oris' or 'Noma'. We highlight the possible association of Pseudomonas sepsis and Noma, with malnutrition playing a central role in causing both the diseases.

Acinetobacter spp as Nosocomial Pathogen : Clinical Significance and Antimicrobial Sensitivity.

Acinetobacter spp are ubiquitous aerobic Gram negative coccobacillus, that are now increasingly responsible for a large number of nosocomial infections. In our study, over a period of six months (Jan-Jun 2000) at a tertiary care hospital, 152 (12.9%) isolates of Acinetobacter spp were obtained from a total of 1175 isolates grown from all clinical specimens. Most of the isolates 126 (82.9%) were from hospitalised patients in the spinal cord injury centre, intensive care units and those on prior antibiotic therapy. Community acquired infections were also seen in 26 (17.1%) out patient department (OPD) cases. Isolates were from urine, respiratory exudates, blood and pus/burn wound swabs predominantly. They were resistant to commonly used antibiotics while being sensitive to amikacin, augmentin, piperacillin, netilmicin and cefotaxime. 69.2% isolates exhibited resistance to two or more antibiotics. Clinical co-relation must be under taken to exclude commensal contaminants, before considering it to be a pathogen and prescribing antibiotics to the patient.

A new approach for modification of phenylalanine peptides by Suzuki-Miyaura coupling reaction.

For the first time, we have modified phenylalanine peptides by the Suzuki-Miyaura coupling reaction which may be useful in developing combinatorial libraries of peptidomimetics.