Clinical study of extrapulmonary head and neck tuberculosis in an urban setting.

Tuberculosis (TB) of the head and neck region is quite common in endemic countries, but is still misdiagnosed due to its varied presentation and different sites of involvement. The aims of the present study were to present the diversities of presentation of head and neck tuberculosis with the diagnostic predicaments faced during evaluation and to assess treatment response to anti-tubercular treatment (ATT). We analysed 48 patients with head and neck tuberculosis who presented to the Department of Otorhinolaryngology in our tertiary care urban hospital over a period of two years from 2013 to 2015 and recorded their data, which included presenting complaints, local and systemic examination findings, investigation results and treatment outcomes. The results showed that majority (64.5%) of cases were female and none of the patients were HIV positive. The most common manifestation was cervical lymphadenopathy (81.25%) with level II being the most commonly affected (31.3%). Three of the 48 patients had coexisting pulmonary TB. Fine needle aspiration cytology (FNAC), histopathological diagnosis and acid fast bacilli (AFB) staining were used to confirm diagnosis. All patients were treated with Category I ATT, which achieved cure in 96.8% of cases. Though cervical lymphadenitis is the most common presentation of head and neck TB, isolated involvement of the sinonasal region, larynx, oral cavity and other sub-sites are not solely unknown entities. It is, therefore, important for clinicians to be aware of atypical and misleading presentations and consider TB as a major differential diagnosis in the head and neck region, even in non-immunocompromised individuals.

ß-Catenin is required for Ron receptor-induced mammary tumorigenesis.

Our previous studies demonstrated that selective overexpression of the Ron receptor tyrosine kinase in the murine mammary epithelium leads to mammary tumor formation. Biochemical analysis of mammary tumor lysates showed that Ron overexpression was associated with increases in ß-catenin expression and tyrosine phosphorylation. ß-Catenin has also been shown to be regulated through tyrosine phosphorylation by the receptor tyrosine kinases Met, Fer and Fyn. However, the molecular and physiological roles of ß-catenin and ß-catenin tyrosine phosphorylation downstream of Ron are not known. To investigate this association, we show that Ron and ß-catenin are coordinately elevated in human breast cancers. Our data also demonstrate that activation of Ron, through ligand binding by hepatocyte growth factor-like protein (HGFL), induces the tyrosine phosphorylation of ß-catenin, primarily on tyrosine residues Tyr 654 and Tyr 670. In addition, HGFL-mediated Ron activation induces both ß-catenin nuclear localization and transcriptional activity, with Tyr 654 and Tyr 670 residues of ß-catenin being critical for these processes. We also demonstrate that a knockdown of Ron in breast cancer cell lines leads to a loss of HGFL-induced ß-catenin-dependent transcriptional activation and cell growth, which can be rescued by activation of canonical Wnt/ß-catenin signaling. Moreover, we show that HGFL-dependent Ron activation mediates upregulation of the ß-catenin target genes cyclin D1 and c-myc, and that expression of these target genes in breast cancer cells is decreased following inhibition of Ron and/or ß-catenin. Finally, we show that genetic ablation of ß-catenin in Ron-expressing breast cancer cells decreases cellular proliferation in vitro, as well as mammary tumor growth and metastasis, following orthotopic transplantation into the mammary fat pad. Together, our data suggest that ß-catenin is a crucial downstream regulator of Ron receptor activation and is an important mediator of mammary tumorigenesis.

Comparison of epilenticular IOL implantation vs technique of anterior and primary posterior capsulorhexis with anterior vitrectomy in paediatric cataract surgery.

PURPOSE: To compare the functional outcome of epilenticular intraocular lens (IOL) implantation vs the technique of anterior continuous curvilinear capsulorhexis (ACCC), posterior continuous curvilinear capsulorhexis (PCCC) with vitrectomy and in-the-bag IOL implantation in paediatric cataract surgery. METHODS: Forty eyes of 33 children with developmental or traumatic cataract, whose mean age was 2-12 years, were randomly divided into two groups A and B. Group A patients underwent epilenticular IOL implantation while in group B patients, ACCC, PCCC with anterior vitrectomy with in-the-bag IOL implantation was performed. Equal number of eyes (10 each) with developmental cataracts (subgroups A1 and B1) and traumatic cataracts (subgroups A2 and B2) were allotted to both the groups. Postoperative visual acuity, opacification of the visual axis, and possible complications were observed and analysed. RESULTS: Four eyes in subgroup B2 had fibrous or ruptured capsules, and were managed by epilenticular IOL implantation technique. One eye in subgroup B2 developed central posterior capsular opacification and hence required a secondary capsulotomy. All cases in group A maintained a clear visual axis at the last follow-up. Minimal postoperative inflammation was noticed in all groups, which subsided with anti-inflammatory medication. At the last follow-up, all eyes in group A gained visual acuity >/=6/18. Whereas in group B, visual acuity >/=6/18 was obtained in 85.7% cases with the epilenticular IOL implantation technique and in 83.3% cases with ACCC and PCCC with anterior vitrectomy technique. CONCLUSION: Epilenticular IOL implantation offers a safe and effective alternative for management of paediatric cataract. In selected cases of traumatic cataract, it is the preferred treatment modality.

Emphysema in heavy smokers with normal chest radiography. Detection and quantification by HCRT.

PURPOSE: To determine the severity and extent of emphysema in heavy smokers by high-resolution CT (HRCT) and to correlate the findings with spirometric tests (STs) and symptomatology. MATERIAL AND METHODS: Fifty adult smokers with a mean age of 53 years with a smoking history of more than 30 pack years and normal chest radiographs underwent HRCT of the chest and ST (FEV1, FEV1/FVC, PEFR). Among these, 22 had symptoms of pulmonary disease and 28 were asymptomatic. Quantification of emphysema was done using a density mask program and the visual scoring method. The results were correlated with ST and symptomatology. RESULTS: 58% (29 out of 50) of the subjects had significant emphysema on HRCT. Eleven out of 15 with normal ST showed emphysema on HRCT while 2 with airflow obstruction on ST showed normal CT scores. 14% (4 out of 28) asymptomatic subjects had severe emphysema compared to 64% of symptomatic subjects. Emphysematous changes were predominantly seen in upper lung zones in 48% of the patients while in 52% it was distributed equally in both upper and lower zones. The number of pack years of smoking showed a positive correlation with CT scores. The correlation between HRCT scores and ST was statistically significant. CONCLUSION: A significant number of asymptomatic and clinically undiagnosed smokers tend to have significant emphysema. HRCT helps in early detection of disease and thus helps implementation of preventive measures.

Itih-4, a serine protease inhibitor regulated in interleukin-6-dependent liver formation: role in liver development and regeneration.

Inter-alpha-trypsin inhibitor-4 (Itih-4) is a liver-restricted member of the serine protease inhibitor family with diverse functions as an anti-apoptotic and matrix stabilizing molecule that are important throughout development. We investigate the functional role of Itih-4 in liver formation, regeneration (LR) and examine its role in calcium and hyaluronic acid binding. Itih-4 expression is prominent in early liver development at E9 and later at E16, being restricted to hepatoblasts, immature hepatocytes, and differentiated hepatocytes. We note a marked and differential increase in Itih-4 labeling in proliferating hepatocytes, compared with bile duct cells in liver explant cultures treated with interleukin-6 (IL-6). After partial hepatectomy, maximal Itih-4 expression occurs in a bimodal manner at 30 min and at 12 hr, with a predominant centrizonal distribution. There is no detectable binding of glutathione transferase-fusion Itih-4 protein to calcium and hyaluronic acid, indicating a possible requirement for posttranslational modifications for these functions. These results suggest that in LR, Itih-4 expression corresponds to that of immediate early genes and may contribute to the entry of normally quiescent hepatocytes into the early stages of the cell cycle. The markedly high expression of Itih-4 in early liver development and in explants treated with IL-6 suggests a prominent role for Itih-4 at key points in liver formation.

The processing and utilization of hepatocyte growth factor/scatter factor following partial hepatectomy in the rat.

Hepatocyte growth factor/scatter factor (HGF/SF) is a pluripotent growth factor capable of acting as a motogen, a morphogen, and a mitogen. Originally, HGF/SF was found as a blood-borne mitogen for hepatocytes and has since been determined to be very important in liver repair. Previous studies have established that HGF/SF must be proteolytically cleaved to elicit its effects. After liver injury by toxins such as carbon tetrachloride or after surgical resection, partial hepatectomy (PHX), HGF/SF concentrations increase in the blood. The aims of this study were to examine (1) which form of HGF/SF is present in the normal liver, (2) which form is present in the regenerating liver after PHX, and (3) if the HGF/SF used after PHX is derived from existing liver reservoirs. Both single-chain HGF/SF and active two-chain HGF/SF are present in normal liver, with the former being the dominant form. After PHX, the liver can be described as having two phases with regard to the use of endogenous HGF/SF. The first phase from 0 to 3 hours is the consumptive phase and is characterized by a decrease in both single-chain HGF/SF and active two-chain HGF/SF. The second phase is the productive phase. It is characterized by a pronounced reappearance of both single-chain HGF/SF as well as two-chain HGF/SF. The activation index shows a 5-fold increase over sham operations during the productive phase. The use of radiolabeled HGF/SF showed that during the first 3 hours, HGF/SF is used in part from hepatic stores. Furthermore, during the first 3 hours after PHX, only active two-chain HGF/SF is seen in the plasma.

Smad proteins and hepatocyte growth factor control parallel regulatory pathways that converge on beta1-integrin to promote normal liver development.

Smads serve as intracellular mediators of transforming growth factor beta (TGF-beta) signaling. After phosphorylation by activated type I TGF-beta receptors, Smad proteins translocate to the nucleus, where they serve as transcription factors and increase or decrease expression of TGF-beta target genes. Mice lacking one copy each of Smad2 and Smad3 suffered midgestation lethality due to liver hypoplasia and anemia, suggesting essential dosage requirements of TGF-beta signal components. This is likely due to abnormal adhesive properties of the mutant hepatocytes, which may result from a decrease in the level of the beta1-integrin and abnormal processing and localization of E-cadherin. Culture of mutant livers in vitro revealed the existence of a parallel developmental pathway mediated by hepatocyte growth factor (HGF), which could rescue the mutant phenotype independent of Smad activation. These pathways merge at the beta1-integrin, the level of which was increased by HGF in the cultured mutant livers. HGF treatment reversed the defects in cell proliferation and hepatic architecture in the Smad2(+/-); Smad3(+/-) livers.

Changes in WNT/beta-catenin pathway during regulated growth in rat liver regeneration.

The wnt/beta-catenin pathway is important during embryogenesis and carcinogenesis. beta-Catenin interaction with E-cadherin has been shown to be crucial in cell-cell adhesion. We report novel findings in the wnt pathway during rat liver regeneration after 70% partial hepatectomy using Western blot analyses, immunoprecipitation studies, and immunofluorescence. We found wnt-1 and beta-catenin proteins to be predominantly localized in hepatocytes. Immediately following partial hepatectomy, we observed an initial increase in beta-catenin protein during the first 5 minutes with its translocation to the nucleus. We show this increase to be the result of decreased degradation of beta-catenin (decrease in serine phosphorylated beta-catenin) as seen by immunoprecipitation studies. We observed activation of beta-catenin degradation complex comprising of adenomatous polyposis coli gene product (APC) and serine-phosphorylated axin protein, beginning at 5 minutes after hepatectomy, leading to its decreased levels after this time. Quantitative changes observed in E-cadherin protein during liver regeneration are, in general, reverse to those seen in beta-catenin. In addition, using immunoprecipitation, we observe elevated levels of tyrosine-phosphorylated beta-catenin at 6 hours onward. Thus, changes in the wnt pathway during regulated growth seem to tightly regulate cytosolic beta-catenin levels and may be contributing to induce cell proliferation and target gene expression. Furthermore, these changes might also be intended to negatively regulate cell-cell adhesion for structural reorganization during the process of liver regeneration.

Expansion of hepatic and hematopoietic stem cells utilizing mouse embryonic liver explants.

Ex vivo embryonic liver explant culture is a novel and attractive approach to obtain abundant hepatic and hematopoietic stem cells. Gene therapy of autologous hepatic and hematopoietic stem cells represents an alternative therapeutic approach to liver transplantation for genetic and metabolic disorders. In this study we characterize the growth and differentiation of hepatic stem cells utilizing embryonic liver cultures. Day 9.5 liver buds are microdissected and cultured under specific conditions. Modulation of growth conditions by addition of hepatocyte growth factor, Flt-3 ligand, and stem cell factor leads to enrichment of hepatic progenitor cells in embryonic liver explants. Under these conditions, we also demonstrate the role of a novel marker PRAJA-1 to identify hepatic stem cells and transitional hepatocytes. Utilization of dexamethasone enhanced pseudolobule formation with increased hepatocytic and biliary differentiation. Transforming growth factor-beta leads to enrichment of biliary cells in the culture. Gut formation is enhanced in the presence of interleukin-3 and blood formation by increasing the mesodermal tissue in these cultures. We also show increased retroviral-mediated expression of the green fluorescent protein expression in the expanded hepatic and hematopoietic stem cells under different culture conditions. Thus, the embryonic liver explant culture is an attractive source for hepatic progenitors and is a possible step towards generating nontumorigenic immortalized hepatocytes with possible transplantation applications.

Screen for Child Anxiety-Related Emotional Disorders (SCARED): convergent and divergent validity.

The purpose of this study was to examine the Screen for Child Anxiety-Related Emotional Disorders' (SCARED) divergent and convergent validity and its ability to identify anxious children. The SCARED, the Child Behaviour Checklist (CBCL), and the State-Trait Anxiety Inventory for Children (STAIC) were administered to children, adolescents (n = 295), and their parents attending an outpatient mood and anxiety disorders clinic. DSM-IIIR/IV diagnoses were made using a semistructured interview (n = 130) or a symptom checklist (n = 165). The Multi-Trait Multi-Method Matrix was used to assess construct validity, and Receiver Operating Curve analysis was used to assess the sensitivity and specificity of the SCARED, CBCL, and STAIC. The SCARED correlated significantly better with the CBCL's internalizing factors than with the externalizing factors. In addition, parent and child forms of the SCARED correlated significantly with the trait and state subscales of the STAIC. Children with an anxiety disorder scored significantly higher on the SCARED than children with depression only or disruptive disorders only (P < 0.05), thus demonstrating the discriminant validity of the SCARED. The SCARED is a reliable and valid screening tool for clinically referred children and adolescents with anxiety disorders.

Intratumoral therapy of cisplatin/epinephrine injectable gel for palliation in patients with obstructive esophageal cancer.

Obstructing esophageal cancer produces severe dysphagia with ensuing death within 90 days. Palliation is possible with modalities like stent placement, laser, and photodynamic therapy. However, these treatments have a high rate of complications, and the overall mortality is not altered. A new alternative treatment evaluated in this study is endoscopic intratumoral injection with cisplatin/epinephrine (CDDP/epi) gel. CDDP/epi gel injections were administered weekly for 3 to 8 weeks in nine patients, median age, 72 years; mean tumor volume (+/-SEM), 41.44 (+/-22.4) cm3. Eight patients had stage IV, and one had stage III esophageal carcinoma. The mean dysphagia score (+/-SEM) was 3.5 (+/-0.17). All patients were followed up until death. Dysphagia resolved in eight patients with reduction in mean dysphagia score (+/-SEM) from 3.5 (+/-0.17) to 0.75 (+/-0.28; p = 0.005). Tumor volume was reduced by 75% in one patient and by 50% in two patients. The median survival was 4 months. The longest follow-up has been 15 months (458 days). In this pilot study, intratumoral injection of CDDP/epi gel restored swallowing in eight of nine patients and was an effective and safe outpatient treatment in patients with obstructive esophageal cancer.

Psychometric properties of the Screen for Child Anxiety Related Emotional Disorders (SCARED): a replication study.

OBJECTIVE: To replicate and extend work on the psychometric properties of the Screen for Child Anxiety Related Emotional Disorders (SCARED), a child and parent self-report instrument used to screen for children with anxiety disorders. METHOD: The 41-item version of the SCARED was administered to a new sample of 190 outpatient children and adolescents and 166 parents. The internal consistency, discriminant, and convergent validity were assessed. In addition, using discriminant function analysis, a briefer version of the SCARED was developed. RESULTS: Using item analyses and factor analyses on the 41-item version, 5 factors were obtained: panic/somatic, generalized anxiety, separation anxiety, social phobia, and school phobia. In general, the total score and each of the 5 factors for both the child and parent SCARED demonstrated good internal consistency and discriminant validity (both between anxiety and depressive and disruptive disorders and within anxiety disorders). A reduced version of the SCARED yielded 5 items and showed similar psychometrics to the full SCARED. CONCLUSIONS: In a new sample, the authors replicated their initial psychometric findings that the SCARED is a reliable and valid instrument to screen for childhood anxiety disorders in clinical settings. Furthermore, pending future research, the 5-item SCARED appears to be a promising brief screening inventory for anxiety disorders in epidemiological studies.

Elf3 encodes a novel 200-kD beta-spectrin: role in liver development.

beta-spectrins are crucial for the maintenance of cell shape, the establishment of cell polarity, and the formation of distinct membrane domains. Our strategy for identifying genes important for hepatocyte polarity has been to utilize subtractive hybridization of early embryonic mouse cDNA liver libraries. As a result, we have cloned three isoforms of a novel beta-spectrin elf (embryonic liver beta-fodrin), and here we report the analysis of elf3, the longest isoform (8172 nt). ELF3 comprises 2154 residues with an overall similarity of 89.0% and 95.3% to mouse beta-spectrin (betaSpIIsigma1) at the nucleotide and amino acid level, respectively. ELF3 is characterized by an actin-binding domain, a long repeat domain, and a short regulatory domain remarkable for the absence of a PH domain. Linkage analysis reveals that elf3 maps to mouse chromosome 11 between D11Bir6 and D11Xrf477, a different chromosomal locus from that of the other four spectrin genes. Northern blot analysis utilizing an elf3 3'-UTR probe demonstrates an abundant 9.0-kb transcript in brain, liver, and heart tissues. Western blot with a polyclonal antibody against ELF identifies a 200 kD protein in mouse liver, brain, kidney, and heart tissues. Immunohistochemical studies demonstrate ELF labeling of the basolateral or sinusoidal membranes surface as well as a granular cytoplasmic pattern in hepatocytes. Antisense studies utilizing cultured liver explants show a vital role of elf3 in hepatocyte differentiation and intrahepatic bile duct formation. The differential expression, tissue localization, and functional studies demonstrate the importance of elf3 in modulating interactions between various components of the cytoskeleton proteins controlling liver and bile duct development.

Genomic structure, chromosomal mapping, and muscle-specific expression of a PH domain-associated intronless gene, cded/lior.

The aim of our study was to isolate novel gene(s) involved in cell differentiation and embryonic liver development. Mouse cded/lior was identified from subtraction hybridization of embryonic liver cDNA libraries as well as an adult mouse liver genomic DNA library. The full open reading frame of cded/lior encodes a 131-amino acid protein with 71.88% overall similarity to the PH domain of rat PLC-gamma1. A gapped search with the C-terminal region of CDED/LIOR revealed a 36-41% similarity to several proteins related to signal transduction and cell replication, such as ORC1 and KSR. Northern blot analysis of adult mouse tissues shows a strong 2.6-kb transcript restricted to heart and skeletal muscle. RT-PCR utilizing cded/lior-specific primers demonstrates cded/lior mRNAs in heart, brain, and liver tissue throughout mid-embryonic mouse gestation. cded/lior maps to the distal end of mouse Chromosome (Chr) 2. Analysis of the genomic structure for cded/lior demonstrated a single exon gene that is not an alternatively spliced isoform of PLC-gamma1. Analysis of the cded/lior promoter region revealed a high GC-content, high ratio of CpG/GpC, multiple GC-boxes, the lack of a TATA box, CTF/NFI element, and two MyoD-MCK binding sites. These characteristics are also found in several genes important in the regulation of cell growth or DNA synthesis, such as transforming growth factor-beta1, c-Ha-ras, nerve growth factor, epidermal growth factor receptor, and DNA polymerase beta. These results suggest that cded/lior is a mesoderm/muscle-specific transcript that may be involved in the mesodermal inductive and regulatory interactions required for liver formation and embryonic development.

Identification of mouse itih-4 encoding a glycoprotein with two EF-hand motifs from early embryonic liver.

An essential feature of cell differentiation is the specificity of signal transduction events from extracellular cues, which are considered to be conferred by scaffold, anchoring and adaptor proteins. Our aim was to identify important scaffolding proteins required for liver development. Utilizing subtraction hybridization of embryonic liver cDNA libraries, here we report the full length cDNA sequence for mouse itih-4 (Inter-alpha-trypsin inhibitor H4). Itih-4 encodes a 942 amino acid protein containing two EF-hand (helix-loop-helix) motifs with an unique short loop, with a potential calcium-binding function. Itih-4 is expressed as a strong 3.1-kb transcript in liver, to a lesser extent in lung and heart tissue. RT-PCR demonstrates itih-4 mRNAs abundantly in liver, less in heart and brain, during mid-embryonic gestation. These results suggest that itih-4 is a potential regulator for extracellular matrix proteins and plays a role during early embryonic liver development.

Praja1, a novel gene encoding a RING-H2 motif in mouse development.

As part of a cloning strategy to identify genes involved in early mouse liver development we have isolated Praja1, a gene with similar sequences to the Drosophila melanogaster gene goliath (gl) which is involved in the fate of mesodermal cells ultimately forming gut musculatures, fat body, and the heart. Praja1 is a 2.1 kb gene encoding a putative 396 amino acid ORF and includes a COOH-terminal RING-H2 domain. Using the Jackson Laboratory BSS panel, we have localized Praja1 on chromosome X at 36 cM, which may be a candidate gene for mouse sla (sex linked sideroblastic anemia), near the X inactivation center gene, Xist. Northern blot analysis demonstrated three transcripts (3.1, 2.6 and 2.1 kb) in mRNA from adult mouse tissues brain, liver, and kidney as well as in mRNA from developing mouse embryos (days 7, 11, 15 and 17 post coitus, p.c.). In vitro transcription/translation yielded a product with an Mr of 59 kD. Immunohistochemical staining of in vitro liver explant cultures using a heterologous antibody against praja1 demonstrated cytoplasmic staining of cuboidal cells that have hepatocyte morphology and organization. The presence of the RING-H2 domain, a proline-rich region at the COOH-end, and regions rich in acidic amino acids, leads to the hypothesis that the Praja1 product is possibly involved in mediating protein-protein interactions, possibly as part of a protein sorting or transport pathway. This is strengthened by the similarity of Praja1 to rat Neurodap1, whose product has been shown to localize to the endoplasmic reticulum and golgi in brain.