ANTICANCER ACTIVITY OF PHLORETIN COMPOUND PURIFIED FROM IRAQI MALUS DOMESTICA L. (APPLE) LEAVES.

The present study was dealing with a Polyphenolic compound known as Phloretin. Phloretin (Ph), a dihydrochalcone, was determined qualitatively and quantitatively in different aerial parts for Iraqi Malus domestica (apple), cv." Ibrahimi" included leaves, petioles, stems, fruit pulp, and peels extracts. Leaves represented a rich source of Ph, which was separated and purified by preparative HPLC. The chemical structure of the isolated Phloretin (Ph2) was confirmed using various analytical characterization techniques: TLC, HPLC, FTIR, Melting point, CHN elemental analyses, 1H-NMR, and 13C-NMR). The scavenging efficacy of Ph2 by DPPH assay was employed. Cytotoxic effect was assessed by MTT assay against cancer cell lines including (Hep G2/ human hepatocyte carcinoma, A549/ human lung adenocarcinoma, SW480 / human colon cancer cell, and AGS /adenocarcinoma of the stomach), beside the non-cancerous cell line (HEK 293). About 1.404 g Ph2 was obtained from 18.146 g apple leaves (7.7%). The DPPH and MTT assay results demonstrated that the purified Ph2 possessed potent antioxidant activity with significant anticancer effects on all cancer cell lines. Data suggested that purified Ph2 from Iraqi apple leaves has potential antioxidant, cytotoxicity, which may benefit in human health.

An overview of the taxonomic instability of endangered Golden and Zhobi mahseer in Pakistan / Uma visão geral da instabilidade taxonômica do golden em perigo e do Zhobi mahseer no Paquistão

In South and South East Asia three genera of fish species i.e. Tor, Neolissochilus and Naziritor are commonly known as Mahseer with at least 47 species. Among these 23 belongs to genus Tor, 22 to Neolissochilus and one to Naziritor i.e. Naziritor zhobensis. Recently another species added to genus Naziritor is Naziritor chelynoides in India. Among Tor species Tor putitora (Hamilton) is the most widely distributed Mahseer in Pakistan and other countries of the Indian subcontinent. However, based alone on morphological characters some authors identify the Pakistani counterparts as Tor macrolepis (Heckel), (a species presumed to be found exclusively in the Indus River system) distinct from Tor putitora (a species found in Ganga Brahmaputra River system). In order to resolve this taxonomic ambiguity, present study carried out meristic and morphometric measurements of Mahseer collected from a total of 11 water bodies of Pakistan. Ratios between the morphometric characters were calculated and statistically analyzed using t-test and correlation coefficient. Two species identified as Tor putitora and Naziritor zhobensis were the sole Mahseer inhabitants of Indus system in Pakistan. Tor putitora occurred at all surveyed sites while Nazirtor zhobensis had a distribution range from river Zhob to tributaries of river Gomal the right bank tributaries of River Indus. The study corroborates that there are no unequivocal morphological synapomorphies in any existing populations of both species. The study further demonstrates that head length, a character frequently used in Mahseer taxonomy, is not a good measure for species identification. Finally the present study establishes that Naziritor zhobensis still exists in the water bodies of Pakistan and that golden Mahseer occurring in Indus riverine system of Pakistan is Tor putitora.

No sul e sudeste da Ásia, três gêneros de espécies de peixes, ou seja, Tor, Neolissochilus e Naziritor, são comumente conhecidos como Mahseer com pelo menos 47 espécies. Entre estas, 23 pertencem ao gênero Tor, 22 a Neolissochilus e um a Naziritor, ou seja, Naziritor zhobensis. Recentemente, outra espécie adicionada ao gênero Naziritor é Naziritor chelynoides na Índia. Entre as espécies de Tor, Tor putitora (Hamilton) é o Mahseer mais amplamente distribuído no Paquistão e em outros países do subcontinente indiano. No entanto, com base apenas em caracteres morfológicos, alguns autores identificam as contrapartes paquistanesas como Tor macrolepis (Heckel) (uma espécie que se presume ser encontrada exclusivamente no sistema do rio Indo), distinta de Tor putitora (uma espécie encontrada no sistema do rio Ganga Brahmaputra). A fim de resolver essas ambiguidades taxonômicas, o presente estudo realizou medidas merísticas e morfométricas de Mahseer coletadas em um total de 11 corpos d’água do Paquistão. As razões entre os caracteres morfométricos foram calculadas e analisadas estatisticamente usando o teste t e o coeficiente de correlação. Duas espécies identificadas como Tor putitora e Naziritor zhobensis foram os únicos habitantes Mahseer do sistema fluvial Indo no Paquistão. Tor putitora ocorreu em todos os locais pesquisados, enquanto Nazirtor zhobensis tinha uma faixa de distribuição do rio Zhob aos afluentes do rio Gomal, afluentes da margem direita do rio Indo. O estudo corrobora que não há sinapomorfias morfológicas inequívocas em nenhuma das populações existentes de ambas as espécies. O estudo demonstra ainda que o comprimento da cabeça, um caractere frequentemente usado na taxonomia de Mahseer, não é boa medida para identificação das espécies. Finalmente, o presente estudo estabelece que Naziritor zhobensis ainda existe nos corpos d'água do Paquistão e que o Mahseer dourado ocorrendo no sistema fluvial Indo do Paquistão é Tor putitora.

An overview of the taxonomic instability of endangered Golden and Zhobi mahseer in Pakistan

Abstract In South and South East Asia three genera of fish species i.e. Tor, Neolissochilus and Naziritor are commonly known as Mahseer with at least 47 species. Among these 23 belongs to genus Tor, 22 to Neolissochilus and one to Naziritor i.e. Naziritor zhobensis. Recently another species added to genus Naziritor is Naziritor chelynoides in India. Among Tor species Tor putitora (Hamilton) is the most widely distributed Mahseer in Pakistan and other countries of the Indian subcontinent. However, based alone on morphological characters some authors identify the Pakistani counterparts as Tor macrolepis (Heckel), (a species presumed to be found exclusively in the Indus River system) distinct from Tor putitora (a species found in Ganga Brahmaputra River system). In order to resolve this taxonomic ambiguity, present study carried out meristic and morphometric measurements of Mahseer collected from a total of 11 water bodies of Pakistan. Ratios between the morphometric characters were calculated and statistically analyzed using t-test and correlation coefficient. Two species identified as Tor putitora and Naziritor zhobensis were the sole Mahseer inhabitants of Indus system in Pakistan. Tor putitora occurred at all surveyed sites while Nazirtor zhobensis had a distribution range from river Zhob to tributaries of river Gomal the right bank tributaries of River Indus. The study corroborates that there are no unequivocal morphological synapomorphies in any existing populations of both species. The study further demonstrates that head length, a character frequently used in Mahseer taxonomy, is not a good measure for species identification. Finally the present study establishes that Naziritor zhobensis still exists in the water bodies of Pakistan and that golden Mahseer occurring in Indus riverine system of Pakistan is Tor putitora.

Resumo No sul e sudeste da Ásia, três gêneros de espécies de peixes, ou seja, Tor, Neolissochilus e Naziritor, são comumente conhecidos como Mahseer com pelo menos 47 espécies. Entre estas, 23 pertencem ao gênero Tor, 22 a Neolissochilus e um a Naziritor, ou seja, Naziritor zhobensis. Recentemente, outra espécie adicionada ao gênero Naziritor é Naziritor chelynoides na Índia. Entre as espécies de Tor, Tor putitora (Hamilton) é o Mahseer mais amplamente distribuído no Paquistão e em outros países do subcontinente indiano. No entanto, com base apenas em caracteres morfológicos, alguns autores identificam as contrapartes paquistanesas como Tor macrolepis (Heckel) (uma espécie que se presume ser encontrada exclusivamente no sistema do rio Indo), distinta de Tor putitora (uma espécie encontrada no sistema do rio Ganga Brahmaputra). A fim de resolver essas ambiguidades taxonômicas, o presente estudo realizou medidas merísticas e morfométricas de Mahseer coletadas em um total de 11 corpos dágua do Paquistão. As razões entre os caracteres morfométricos foram calculadas e analisadas estatisticamente usando o teste t e o coeficiente de correlação. Duas espécies identificadas como Tor putitora e Naziritor zhobensis foram os únicos habitantes Mahseer do sistema fluvial Indo no Paquistão. Tor putitora ocorreu em todos os locais pesquisados, enquanto Nazirtor zhobensis tinha uma faixa de distribuição do rio Zhob aos afluentes do rio Gomal, afluentes da margem direita do rio Indo. O estudo corrobora que não há sinapomorfias morfológicas inequívocas em nenhuma das populações existentes de ambas as espécies. O estudo demonstra ainda que o comprimento da cabeça, um caractere frequentemente usado na taxonomia de Mahseer, não é boa medida para identificação das espécies. Finalmente, o presente estudo estabelece que Naziritor zhobensis ainda existe nos corpos dágua do Paquistão e que o Mahseer dourado ocorrendo no sistema fluvial Indo do Paquistão é Tor putitora.

Monitoring pond water quality to improve the production of Labeo rohita (Hamilton, 1822) in Bannu Fish Hatchery of Bannu district, Khyber Pakhtunkhwa province; An Implications for artificial fish culture

Abstract The water quality parameters of a fish pond are essential to be managing properly under control for successful operations of fish culture. Improper management of pond water quality during the juvenile stages can create stressful conditions to produce various harmful diseases, which may decrease the fish quality and results in low profits. The present study was aimed to monitored important water quality parameters of nursery ponds of Labeo rohita culturing in Bannu fish hatchery. The study duration was 75 days extends from 10th June to 24th August 2019 for the successful culture of this specie which can play a significant role in breeding season. Furthermore, the concentration of some heavy metals like copper (Cu), nickel (Ni), manganese (Mn), iron (Fe), cadmium (Cd), and zinc (Zn) in pond water and fry stages of this species was also determined. The data obtained from all water quality parameters were analyzed expressed as range, mean and standard deviation using MS Excel 2013. The obtained results of 75 days study revealed that the water pH & temperature, electric conductivity, total dissolved solids (TDS), and total dissolved oxygen (DO) of pond water samples were found within a tolerable limit except salinity and dissolved ammonia concentration were not permissible for fish growth. The average concentration of heavy metals in pond water exhibited descending order Fe>Ni>Mn>Zn>Cd>Ni, which was found in acceptable ranges. Whereas, the average values of heavy metals in fry stages were in the order of Zn>Fe>Ni>Cu>Cd>Mn, and found within the recommended values of WHO/FAO. Thus, it was concluded from this study that good water quality is a precondition, maintaining balanced levels of water quality parameters is fundamental for both the health and growth of fish culture which is quite necessary for assuring increased fish productivity. It is recommended to monitor and assess water quality parameters on a routine basis for promoting healthy fish culture.

Resumo Os parâmetros de qualidade da água de um viveiro de peixes são essenciais para um manejo adequado e sob controle para operações bem-sucedidas de piscicultura. O manejo inadequado da qualidade da água do tanque durante os estágios juvenis pode criar condições estressantes para a produção de várias doenças nocivas, o que pode diminuir a qualidade do peixe e resultar em baixos lucros. O presente estudo teve como objetivo monitorar importantes parâmetros de qualidade da água de viveiros de cultivo de Labeo rohita em incubadora de peixes Bannu. A duração do estudo foi de 75 dias, estendendo-se de 10 de junho a 24 de agosto de 2019 para o sucesso do cultivo dessa espécie que pode desempenhar papel significativo na época de reprodução. Além disso, a concentração de alguns metais pesados como cobre (Cu), níquel (Ni), manganês (Mn), ferro (Fe), cádmio (Cd) e zinco (Zn) na água do tanque e estágios de fritura dessa espécie também foram determinados. Os dados obtidos de todos os parâmetros de qualidade da água foram analisados expressos como faixa, média e desvio padrão usando o MS Excel 2013. Os resultados obtidos de 75 dias de estudo revelaram que o pH e temperatura da água, condutividade elétrica, sólidos totais dissolvidos (TDS) e total de oxigênio dissolvido (OD) das amostras de água do lago foram encontrados dentro de um limite tolerável, exceto salinidade e concentração de amônia dissolvida não eram permitidas para o crescimento dos peixes. A concentração média de metais pesados na água da lagoa apresentou ordem decrescente Fe > Ni > Mn > Zn > Cd > Ni, que foi encontrada em faixas aceitáveis. Já os valores médios dos metais pesados nos estágios de fritura foram da ordem de Zn > Fe > Ni > Cu > Cd > Mn, e encontrados dentro dos valores recomendados pela OMS/FAO. Assim, concluiu-se deste estudo que a boa qualidade da água é uma condição prévia, manter níveis equilibrados dos parâmetros de qualidade da água é fundamental para a saúde e crescimento da piscicultura, o que é bastante necessário para garantir o aumento da produtividade piscícola. Recomendam-se monitorar e avaliar os parâmetros de qualidade da água em uma base rotineira para promover a piscicultura saudável.

An overview of the taxonomic instability of endangered Golden and Zhobi mahseer in Pakistan / Uma visão geral da instabilidade taxonômica do golden em perigo e do Zhobi mahseer no Paquistão

Abstract In South and South East Asia three genera of fish species i.e. Tor, Neolissochilus and Naziritor are commonly known as Mahseer with at least 47 species. Among these 23 belongs to genus Tor, 22 to Neolissochilus and one to Naziritor i.e. Naziritor zhobensis. Recently another species added to genus Naziritor is Naziritor chelynoides in India. Among Tor species Tor putitora (Hamilton) is the most widely distributed Mahseer in Pakistan and other countries of the Indian subcontinent. However, based alone on morphological characters some authors identify the Pakistani counterparts as Tor macrolepis (Heckel), (a species presumed to be found exclusively in the Indus River system) distinct from Tor putitora (a species found in Ganga Brahmaputra River system). In order to resolve this taxonomic ambiguity, present study carried out meristic and morphometric measurements of Mahseer collected from a total of 11 water bodies of Pakistan. Ratios between the morphometric characters were calculated and statistically analyzed using t-test and correlation coefficient. Two species identified as Tor putitora and Naziritor zhobensis were the sole Mahseer inhabitants of Indus system in Pakistan. Tor putitora occurred at all surveyed sites while Nazirtor zhobensis had a distribution range from river Zhob to tributaries of river Gomal the right bank tributaries of River Indus. The study corroborates that there are no unequivocal morphological synapomorphies in any existing populations of both species. The study further demonstrates that head length, a character frequently used in Mahseer taxonomy, is not a good measure for species identification. Finally the present study establishes that Naziritor zhobensis still exists in the water bodies of Pakistan and that golden Mahseer occurring in Indus riverine system of Pakistan is Tor putitora.

Resumo No sul e sudeste da Ásia, três gêneros de espécies de peixes, ou seja, Tor, Neolissochilus e Naziritor, são comumente conhecidos como Mahseer com pelo menos 47 espécies. Entre estas, 23 pertencem ao gênero Tor, 22 a Neolissochilus e um a Naziritor, ou seja, Naziritor zhobensis. Recentemente, outra espécie adicionada ao gênero Naziritor é Naziritor chelynoides na Índia. Entre as espécies de Tor, Tor putitora (Hamilton) é o Mahseer mais amplamente distribuído no Paquistão e em outros países do subcontinente indiano. No entanto, com base apenas em caracteres morfológicos, alguns autores identificam as contrapartes paquistanesas como Tor macrolepis (Heckel) (uma espécie que se presume ser encontrada exclusivamente no sistema do rio Indo), distinta de Tor putitora (uma espécie encontrada no sistema do rio Ganga Brahmaputra). A fim de resolver essas ambiguidades taxonômicas, o presente estudo realizou medidas merísticas e morfométricas de Mahseer coletadas em um total de 11 corpos d'água do Paquistão. As razões entre os caracteres morfométricos foram calculadas e analisadas estatisticamente usando o teste t e o coeficiente de correlação. Duas espécies identificadas como Tor putitora e Naziritor zhobensis foram os únicos habitantes Mahseer do sistema fluvial Indo no Paquistão. Tor putitora ocorreu em todos os locais pesquisados, enquanto Nazirtor zhobensis tinha uma faixa de distribuição do rio Zhob aos afluentes do rio Gomal, afluentes da margem direita do rio Indo. O estudo corrobora que não há sinapomorfias morfológicas inequívocas em nenhuma das populações existentes de ambas as espécies. O estudo demonstra ainda que o comprimento da cabeça, um caractere frequentemente usado na taxonomia de Mahseer, não é boa medida para identificação das espécies. Finalmente, o presente estudo estabelece que Naziritor zhobensis ainda existe nos corpos d'água do Paquistão e que o Mahseer dourado ocorrendo no sistema fluvial Indo do Paquistão é Tor putitora.

Patient preferences for topical treatment of actinic keratoses: a discrete-choice experiment.

BACKGROUND: The treatment of actinic keratosis (AK) is a potentially effective strategy for the prevention of cutaneous squamous cell carcinoma (cSCC). However, the patient perspective on potential benefits of AK treatment in terms of skin cancer reduction has received little attention to date. OBJECTIVES: (i) To investigate patient preferences for topical treatments for AK using a discrete-choice experiment (DCE); (ii) to evaluate patient willingness to trade between clinical benefit and medical burden. METHODS: The DCE was conducted as part of a study to establish the feasibility of a phase III randomized controlled trial evaluating the prevention of cSCC using currently available topical interventions. Preferences were elicited by asking patients to make a series of choices between treatment alternatives with different hypothetical combinations of attribute levels. Willingness to trade between treatment attributes was estimated using a flexible-choice model that allows for the heterogeneity of patient preferences. RESULTS: A total of 109 patients with AK completed the DCE. The majority of patients who expressed valid preferences were willing to accept some reduction in both prophylactic and cosmetic efficacy to reduce the burden of the treatment regimen, the severity of skin reaction and other adverse effects. Patients may reject treatment if the perceived therapeutic benefit is outweighed by the subjective burden of treatment. CONCLUSIONS: Evidence of significant variation in the perceived utility of treatments across patients highlights the importance of taking individual patient preferences into account to improve AK treatment acceptability and adherence.

Systematic review and meta-analysis of the complications of salvage total laryngectomy.

BACKGROUND AND OBJECTIVES: Management paradigms in laryngeal cancer have shifted to "organ preservation" chemoradiotherapy protocols. In the event of treatment failure, salvage total laryngectomy remains the only curative treatment option. However a comprehensive review of the complications of this procedure has not been reported. METHODS: A systematic review of the literature was performed using keywords "salvage laryngectomy" to retrieve relevant publications between January 2000 and August 2015. RESULTS: Of the 407 articles retrieved from the literature search, 50 studies encompassing 3292 patients were included. Forty-nine studies reported pharyngocutaneous fistula which occurred in 859 patients (pooled incidence 28.9%; 95% confidence intervals 25.5-32.5%). Twenty-four studies reported complications in addition to PCF and these included wound complications (infection, dehiscence and necrosis), dysphagia, bleeding, and pharyngeal and stomal stenosis. CONCLUSIONS: Overall complication rate was 67.5%, Pharyngocutaneous fistula was the commonest complication with a pooled incidence of 28.9%.

SOCS1 gene expression is increased in severe pulmonary tuberculosis.

Suppressors of cytokine signalling (SOCS) molecules inhibit cytokine signalling and may regulate protective immunity in tuberculosis (TB). We investigated the association of SOCS with disease progression in patients with pulmonary TB. For this purpose, we studied peripheral blood mononuclear cells (PBMCs) and T cells from patients with pulmonary TB (TB, n=33) and healthy endemic controls (EC, n=15). Cases were stratified into those with moderately advanced (Mod-PTB) or far advanced disease (Adv-PTB). Interferon-gamma (IFN-γ), SOCS1 and SOCS3 gene expression was determined by RT-PCR. Statistical analysis was performed using the Mann-Whitney test. Levels of IL6 (P=0.018) and IL10 (P=0.013) were found to be elevated in PBMC supernatants from patients with TB as compared with EC. SOCS1 mRNA gene expression in T cells from patients with TB was increased as compared with that of EC (P=0.02). In addition, levels of SOCS1 mRNA transcripts were found to be elevated in PBMCs of Adv-PTB as compared with Mod-PTB (P=0.008) cases. Our data show that raised SOCS1 levels are associated with increased disease severity in TB. As SOCS1 regulates IFN-γ-driven immunity and SOCS1 can be further upregulated by IL6 levels, the increase in SOCS1 in severe disease indicates a mechanism by which mycobacteria impede disease control in TB.

Mediastinal parathyroid adenoma: diagnostic and management challenges.

Primary hyperparathyroidism due to ectopic parathyroid adenomas can pose diagnostic and management challenges, especially when imaging studies have localised the lesions to different sites. We report a case of symptomatic hypercalcaemia due to a mediastinal parathyroid adenoma. Ultrasonography identified a nodule posterior to the right thyroid gland. However, computed tomography and technetium-99m sestamibi scintigraphy revealed an ectopic parathyroid adenoma located in the anterior mediastinum. The adenoma was successfully removed through a median sternotomy. However, postoperatively, the patient developed prolonged symptomatic hypocalcaemia, possibly due to suppression of the normal parathyroid gland function, although the presence of concomitant hungry bone syndrome was possible. The histopathology of the mediastinal mass was consistent with a parathyroid adenoma.

Elevated serum CCL2 concomitant with a reduced mycobacterium-induced response leads to disease dissemination in leprosy.

Mycobacterium leprae and Mycobacterium tuberculosis are successful intracellular pathogens which down regulate host immune responses. T-cell interferon-gamma (IFNgamma) and macrophage tumour necrosis factor-alpha (TNFalpha) activate chemokines such as, C-C chemokine ligand-2 (CCL2) and CCL5, which play a role in granuloma formation. Lepromatous leprosy is characterized by defective granulomas with lowered T-cell- and macrophage-mediated responses. Tuberculosis (TB) can be localized to the lung, whereby discreet granulomas are formed. The role of chemokines in leprosy infections is as yet unclear. We compared chemokine responses in lepromatous leprosy and pulmonary tuberculosis patients. Circulating serum CCL2 was raised while CCL5 was lowered in leprosy, as compared with TB patients and healthy controls. However, both Mycobacterium bovis BCG- (P=0.08) and M. leprae-induced (P=0.05) CCL2 secretion was reduced in leprosy. In leprosy, BCG induced greater CCL2 (P=0.01), TNFalpha (P=0.02) and somewhat greater CCL5 (P=0.08) than M. leprae, while CXCL8 induction was comparable. Overall levels of Mycobacterium-induced CCL2, TNFalpha and CXCL8 were two to threefold lower, and CCL5 was 10-fold lower in leprosy as compared with TB. Reduced inducible CCL2 combined with a lowered TNFalpha response in lepromatous leprosy may contribute to the unrestricted growth and dissemination of mycobacteria found in the disease.

Bromate formation on the non-porous TiO2 photoanode in the photoelectrocatalytic system.

The increasing use of ozone in water disinfection processes has been the focus of considerable concern in regards to inorganic disinfection by product formation of bromate in waters containing bromide. Due to the public health risk caused by the presence of bromate as a suspected carcinogen, attention had been addressed to the conditions under which bromate is formed. In this study, photoanodic bromine generation and bromate (BrO(3)(-)) formation were investigated using a TiO(2) electrode in a photoelectrocatalytic (PEC) treatment process. The separation of anodic and cathodic reactions in the PEC system resulted in a pH decrease from 9.3 to 3.0 in the photoanode compartment and an increase to 11.0 in the cathode compartment. Under a photo-illumination intensity of 5.7 m W cm(-2) UV, a biasing potential of +1.0V vs SCE, a pH of 6.0 and at a NaBr concentration of 1.0 x 10(-2) M, active bromine formation increased over time with 2.4 x 10(-6) M min(-6) rate and reached a steady-state concentration of 1.44 x 10(-4) M in 60 min. Bromate formation was detected after a lag-period of 15 min and exhibited a continuous increasing trend with respect to irradiation time. No bromate formation was observed below pH 6.5 whereas an increasing bromate concentrations and pH up to pH=8.5 were noted.

Leprosy patients with lepromatous disease have an up-regulated IL-8 response that is unlinked to TNF-alpha responses.

Tumor necrosis factor (TNF-alpha) in conjunction with interferon-gamma (IFN-gamma) plays an important role in lymphocyte recruitment and granuloma formation in mycobacterial diseases. Lepromatous leprosy infections are typically associated with low to absent T cell responses and the absence of INF-gamma secretion. Chemokines such as IL-8, MCP-1, and MIP-1beta, have also been shown to recruit neutrophils and lymphocytes to the site of mycobacterial infections. We have studied IL-8 expression in relation to TNF-alpha and TGF-beta in monocytes from lepromatous patients (LL) as compared with healthy endemic controls. In endemic controls, no spontaneous expression of IL-8, TNF-alpha, and TGF-beta was observed, but BCG and M. leprae induced activation of all three cytokines. Lepromatous leprosy monocytes spontaneously expressed high levels of IL-8 and TGF-beta but negligible levels of TNF-alpha. A further increase in IL-8 secretion or gene expression by BCG or M. leprae was not significant. BCG, but not M. leprae, was able to stimulate TNF-alpha activation in lepromatous leprosy subjects. TGF-beta responses in LL were parallel to those of IL-8. This suggests a vigorous and active ongoing IL-8 response in lepromatous disease that is independent of TNF-alpha activation. Therefore, in the absence of IFN-gamma and TNF-alpha activation, IL-8 may assume a pivotal role in cell recruitment in leprosy patients with disseminated mycobacterial infections.

IgG1 antimycobacterial antibodies can reverse the inhibitory effect of pentoxifylline on tumour necrosis factor alpha (TNF-alpha) secreted by mycobacterial antigen-stimulated adherent cells.

Chronic inflammation associated with cachexia, weight loss, fever and arthralgia is the hallmark of advanced mycobacterial diseases. These symptoms are attributed to the chronic stimulation of tumour necrosis factor (TNF)-alpha. Mycobacterial components directly stimulate adherent cells to secrete TNF-alpha. We have shown recently that IgG1 antimycobacterial antibodies play a role in augmenting TNF-alpha in purified protein derivative (PPD)-stimulated adherent cells from non-BCG-vaccinated donors. We now show that IgG1 antibodies can also augment TNF-alpha expression in stimulated adherent cells obtained from BCG-vaccinated donors and this augmentation is not linked to interleukin (IL)-10 secretion. In addition IgG1 antimycobacterial antibodies can reverse the effect of TNF-alpha blockers such as pentoxifylline and thalidomide. These studies therefore have clinical implications for anti-inflammatory drug treatments which are used increasingly to alleviate symptoms associated with chronic inflammation.

The effect of mycobacterial virulence and viability on MAP kinase signalling and TNF alpha production by human monocytes.

SETTING: The success of Mycobacterium tuberculosis as a human pathogen depends on its ability to tolerate and perhaps manipulate host defense mechanisms. OBJECTIVE: To determine the induction of tumour necrosis factor-alpha (TNF alpha), a central mediator of immunity, by human monocytes infected with virulent M. tuberculosis, M. leprae and attenuated M. bovis BCG. DESIGN: Mycobacteria-induced cellular activation pathways of TNF alpha production was investigated using an inhibitor of protein tyrosine kinase (PTKs) and an inhibitor of mitogen-activated protein (MAP) kinases. RESULTS: TNF alpha production was significantly lower during infection with virulent M. tuberculosis than with BCG and this differential response was independent of mycobacterial viability. TNF alpha production involved the PTK and MAP kinase pathways. Reduced TNF alpha induction by M. tuberculosis was associated with a reduction in the extent and duration of phosphorylation of extracellular-signal regulated kinases (ERK 1/2). Infection with M. leprae triggered low and transient ERK 1/2 activation as well as low TNF alpha production. CONCLUSION: Maintenance of the differential response in both live and heat-killed preparations suggests that the reduced TNF alpha response associated with virulent mycobacteria is due to differences in the presence of components capable of triggering host pattern recognition receptors, rather than events associated with phagosome trafficking or the active release of intracellular modulators.

The clinical role of interleukin-6 and interleukin-6 soluble receptor in human follicular fluids.

In order to investigate the role of interleukin-6 (IL-6) and interleukin-6 soluble receptor (sR) in human ovulation, we evaluated the concentrations in human follicular fluid and analyzed the correlation of IL-6 and IL-6 sR with oocyte maturation. The oocytes were obtained from the follicular fluid of 45 women undergoing in vitro fertilization and embryo transfer. The concentrations of IL-6 and IL-6 sR in follicular fluid were measured by ELISA. In addition, granulosa cells obtained from the follicular fluid were cultured and treated with forskolin and 12- o-tetradecanoylphorbol 13-acetate for 24-48 h. The concentration of IL-6 was significantly higher in the follicular fluid than in the serum (P<0.01). In contrast, the concentration of IL-6 sR was significantly lower in the follicular fluid than in the serum (P<0.001). The concentrations of IL-6 and IL-6 sR were significantly higher in the follicular fluid containing mature oocytes than in fluid containing immature oocytes (P<0.05). The production of IL-6 was markedly increased over the basal level after 24 h of treatment with forskolin (P<0.001) and 48 h of treatment (P<0.01) with cultured granulosa cells. Our data suggest that IL-6 and IL-6 sR may play an important role in follicular growth and development in human preovulatory processes. It is possible that IL-6 in particular may be regulated by cAMP. IL-6 and IL-6 sR might also be valuable biochemical markers in the evaluation of oocyte maturation.

Assessment of immunity to mycobacterial infection with luciferase reporter constructs.

Protective immunity to mycobacterial infection is incompletely understood but probably involves the coordinated interaction of multiple cell types and cytokines. With the aim of developing assays that might provide a surrogate measure of protective immunity, we have investigated the use of recombinant mycobacteria carrying luciferase reporter enzymes to assess the effectiveness of antimycobacterial immunity in model systems. Measurement of luminescence was shown to provide a rapid and simple alternative to the counting of CFU as a means of monitoring mycobacterial viability. We describe optimization of a luciferase reporter strain of Mycobacterium tuberculosis and demonstrate its application for the study of mycobacterial interactions with host cells in tissue culture and the rapid assessment of vaccine efficacy in a murine model.

Competitive polymerase chain reaction for the determination of N-myc amplification in neuroblastoma: report of clinical cases.

If an unfavorable prognosis is suspected in neuroblastoma, decision on a treatment protocol should be based on the N-myc copy number (12). We already demonstrated that the newly developed competitive polymerase chain reaction (competitive PCR) is a promising method for the determination of the N-myc copy number (6), and have started to use this competitive PCR procedure in neuroblastoma patients, together with fine-needle biopsy in selected cases. Seven children were studied. In one infant of 5 months of age whose tumor was diagnosed before undergoing mass screening for neuroblastoma, the competitive PCR procedure was performed with a fine-needle biopsy, and after obtaining a negative report on N-myc amplification within 48 hours, a regular protocol of treatment could be started without delay. We report that competitive PCR is a rapid and accurate method for the determination of the N-myc copy number, requiring only a small amount of material, and anticipate that competitive PCR will become the procedure of choice for the determination of N-myc copy number in neuroblastoma.

The spinal curve in standing and sitting postures in children with idiopathic scoliosis.

STUDY DESIGN: A sample of convenience of children with moderate idiopathic scoliosis without bracing or surgery was studied. The sample consisted of 19 children, aged 9 to 16 years, with mean Cobb angle of 24 degrees. The spinal configurations and paraspinal muscle activity in several commonly assumed postures were examined. OBJECTIVES: To determine how the apex angles, verticality of spine, and muscle activity vary with the assumed posture and whether the location and the number of spinal curves affect these variables. SUMMARY OF BACKGROUND DATA: It has been suggested that the configuration of the spine in commonly assumed postures can affect the spinal curve in scoliosis because of gravitational bending moments. There is, however, a paucity of data obtained in subjects in sitting postures that school-age children assume daily for prolonged periods. Absence of bilateral symmetry in pressure during sitting has been described, but its effect on the spinal apex angle has not been investigated. METHODS: Infrared-emitting markers, whose three-dimensional positions could be tracked by a pair of cameras, were affixed to the spine. The natural postures studied were relaxed standing, relaxed sitting, erect sitting, and writing while seated. Electromyographic activity in muscles close to the spinal apexes was recorded bilaterally for each test posture. RESULTS: Subjects leaned laterally and, in general, anteriorly, in all test postures, rather than placing the C7 vertebra vertically above S1. The direction of lean and the change in the spinal apex angle from standing to sitting varied depending on whether the spinal curve was single or double, thoracic or lumbar. Subjects with single curves, whether thoracic or lumbar, tended to lean laterally toward the convexity of their curve apex--that is, the lean was in a direction that reduced the apex angle. Subjects with double curves (thoracic and lumbar), in all postures except relaxed sitting, tended to lean toward the convexity of the lumbar curve, thereby reducing the lumbar apex angle and exacerbating the thoracic angle. Most subjects' apex angles were smaller in relaxed or erect sitting than in relaxed standing. Electromyographic activity was in general greater on the convex side of the curve, with greatest activity in erect sitting. CONCLUSIONS: The findings indicate that in self-selected postures the gravitational effect of leaning and the muscle activity in paraspinal muscles may serve to reduce the apex angle. Thus, a fully upright, centered posture may not be best for correction of every patient's spinal curve.

Subcellular fractionation by organelle electrophoresis: separation of phagosomes containing heat-killed yeast particles.

Uptake of foreign material and its subsequent lysosomal degradation is an important function of macrophages. The mechanisms involved in the binding, uptake and delivery of such material to lysosomal organelles, are, however, poorly understood. Here we describe a method using organelle electrophoresis to study the uptake and trafficking of heat-killed yeast particles within murine macrophages. Such yeast particles, which were fluorescently labeled, could be readily detected in intact cells as well as in subcellular fractions. Organelle electrophoresis of a homogenate from macrophages that had internalized yeast particles resulted in the separation of yeast-containing organelles from most other subcellular membranes. In addition, this method was used to follow the kinetics of yeast particle transport within macrophages could be followed readily. Organelle electrophoresis may be a valuable tool for the analysis of phagocytosis.