The human stress-activated protein kinase-interacting 1 gene encodes JNK-binding proteins

The orthologous proteins of the stress-activated protein kinase-interacting 1 (Sin1) family have been implicated in several different signal transduction pathways. In this study, we have investigated the function of the full-length human Sin1 protein and a C-terminally truncated isoform, Sin 1 alpha, which is produced by alternative splicing. Immunoblot analysis using an anti-Sin 1 polyclonal antibody showed that full-length Sin I and several smaller isoforms are widely expressed. Sin 1 was demonstrated to bind to c-Jun N-terminal kinase (JNK) in vitro and in vivo, while no interaction with p38- or ERK1/2-family MAPKs was observed. The Sin1 alpha isoform could also form a complex with JNK in vivo. Despite localizing in distinct compartments within the cell, both Sin1 and Sin1 alpha co-localized with JNK, suggesting that the Sin1 proteins could recruit JNK. Over-expression of full-length Sin1 inhibited the activation of JNK by UV-C in DG75 cells, as well as basal JNK-activity in HEK293 cells. These data suggest that the human Sin1 proteins may act as scaffold molecules in the regulation of signaling by JNK. (c) 2004 Elsevier Inc. All rights reserved.

Genetic control of adventitious rooting on stem cuttings in two Pinus elliottii x P-caribaea hybrid families

Genetic control of adventitious rooting was characterised in two unrelated Pinus elliottii x P. caribaea families, an outbred F-1 (n = 287) and an inbred F-2 ( n = 357). Rooting percentage was assessed in three settings and root biomass was measured on a sub-set of clones ( n = 50) from each family in the third setting. On average, clones in the outbred F-1 had a higher rooting percentage (mean +/- SE; 59 +/- 1.9%) and biomass (mean +/- SD; 0.41 +/- 0.24 g) than clones in the inbred F-2 family ( mean +/- SE; 48 +/- 1.8% and mean +/- SD; 0.19 +/- 0.13 g). Genetic determination for rooting percentage was strong in both families, as indicated by high individual setting clonal repeatabilities ( e. g. Setting 3; outbred F-1 0.62 +/- 0.03 and inbred F-2 0.68 +/- 0.02 (H-2 +/- SE)) and the moderate-to-high genetic correlations amongst the three settings. For root biomass, clonal repeatabilities for both families were lower (outbred F-1 0.35 +/- 0.09 and inbred F-2 0.44 +/- 0.10 (H-2 +/- SE)). Weak positive genetic correlations between rooting percentage and root biomass in both families suggested a concomitant gain in root biomass would be insignificant when selecting solely on the more easily assessable rooting percentage.

Effectiveness of a mutual support group for families of patients with schizophrenia

Aim. This paper reports a study to examine the effectiveness of a 12-session mutual support group for Chinese families caring for a relative with schizophrenia compared with a psycho-educational group and routine family support services in Hong Kong. Background. Schizophrenia is a disruptive and distressing illness for patients and their families. With the current trend of community care for mental illness, there is evidence that family intervention reduces patient relapse and re-hospitalization, satisfies the health needs of families and enhances their coping capabilities. Methods. A randomized controlled trial was conducted from May 2002 to June 2003 with 96 Chinese families of a relative with schizophrenia selected from two psychiatric outpatient clinics in Hong Kong. Families were randomly assigned to receive mutual support (n = 32), psycho-education (n = 33) or standard care only (n = 31). The interventions were delivered at outpatient clinics over a 6-month period. Pre- and post- (1 week and 6 months) testing took place and families' functioning, mental health service utilization, patients' level of functioning and duration of re-hospitalization were measured. Results. At both post-test periods, family caregivers and patients in the mutual support group reported statistically significant improvements on family and patients' level of functioning, when compared with their counterparts in the psycho-education and standard care groups. Conclusions. The findings support the use of mutual support groups as an effective modality of family intervention in a Chinese population caring for a family member with schizophrenia to improve both family and patient functioning.

Intrauterine growth restriction due to uteroplacental vascular insufficiency leads to increased hypoxia-induced cerebral apoptosis in newborn piglets

Uteroplacental vascular insufficiency in humans is a common cause of intrauterine growth restriction (IUGR) and is associated with an increased incidence of perinatal asphyxia and neurodevelopmental disorders compared to normal weight newborns. Experimental models that provide an opportunity to analyze the pathogenesis of these relationships are limited. Here, we used neonatal pigs from large litters in which there were piglets of normal birth weight (for controls) and of low birth weight (for uteroplacental vascular insufficiency). Hypoxia was induced in paired littermates by reducing the fraction of inspired oxygen to 4% for 25 min. Brain tissue was collected 4 h post-hypoxia. Cerebral levels of apoptosis were quantified morphologically and verified with caspase-3 activity and TUNEL. Expression of Bcl-2, BcI-XL and Bax proteins was investigated using immunohistochemistry. Cellular positivity for Bcl-2 was consistently higher in the non-apoptotic white matter of the hypoxic IUGR animals compared with their littermates and reached significance at P < 0.05 in several pairs of littermates. Alterations in Bax showed a trend towards higher expression in the hypoxic IUGR littermates but rarely reached significance. The IUGR piglets showed a significantly greater amount of apoptosis in response to the hypoxia than the normal weight piglets, suggesting an increased vulnerability to apoptosis in the IUGR piglets. (c) 2006 Elsevier B.V. All rights reserved.

Congruence in QTL for adventitious rooting in Pinus elliottii X Pinus caribaea hybrids resolves between and within-species effects

Targeting between-species effects for improvement in synthetic hybrid populations derived from outcrossing parental tree species may be one way to increase the efficacy and predictability of hybrid breeding. We present a comparative analysis of the quantitative trait loci (QTL) which resolved between from within-species effects for adventitious rooting in two populations of hybrids between Pinus elliottii and P. caribaea, an outbred F-1 (n=287) and an inbred-like F-2 family (n=357). Most small to moderate effect QTL (each explaining 2-5% of phenotypic variation, PV) were congruent (3 out of 4 QTL in each family) and therefore considered within-species effects as they segregated in both families. A single large effect QTL (40% PV) was detected uniquely in the F-2 family and assumed to be due to a between-species effect, resulting from a genetic locus with contrasting alleles in each parental species. Oligogenic as opposed to polygenic architecture was supported in both families (60% and 20% PV explained by 4 QTL in the F-2 and F-1 respectively). The importance of adventitious rooting for adaptation to survive water-logged environments was thought in part to explain oligogenic architecture of what is believed to be a complex trait controlled by many hundreds of genes.

Neural progenitor number is regulated by nuclear factor-kappa B p65 and p50 subunit-dependent proliferation rather than cell survival

The number of cells generated by a proliferating stem or precursor cell can be influenced both by proliferation and by the degree of cell death/survival of the progeny generated. In this study, the extent to which cell survival controls progenitor number was examined by comparing the growth characteristics of neurosphere cultures derived from mice lacking genes for the death inducing Bcl-2 homologue Hara Kiri (Hrk), apoptosis-associated protein 1 (Apaf1), or the prosurvival nuclear factor-kappa B (NF kappa B) subunits p65, p50, or c-rel. We found no evidence that Hrk or Apaf1, and by inference the mitochondrial cell death pathway, are involved in regulating the number of neurosphere-derived progeny. However, we identified the p65p50 NF kappa B dimer as being required for the normal growth and expansion of neurosphere cultures. Genetic loss of both p65 and p50 NF kappa B subunits resulted in a reduced number of progeny but an increased proportion of neurons. No effect on cell survival was observed. This suggests that the number and fate of neural progenitor cells are more strongly regulated by cell cycle control than survival. (c) 2005 Wiley-Liss, Inc.

Ryk: A novel Wnt receptor regulating axon pathfinding

Members of the Wnt family and their receptors, the Frizzleds, are key regulators of pivotal developmental processes including embryonic patterning, specification of cell fate, and determination of cell polarity. The versatility and complexity of Wnt signaling has been further highlighted by the emergence of a novel family of Wnt receptors, the Ryk family. In mammals and flies, Ryk is a key chemorepulsive axon guidance receptor responsible for the establishment of important axon tracts during nervous system development. Although the function of Ryk is currently best understood with respect to this role, its widespread expression, both in developing tissues and in the adult, suggests that Ryk may regulate many essential biological processes. This hypothesis is supported by the multiple developmental phenotypes apparent in Ryk loss-of-function mice. These mice display a variety of embryonic abnormalities, including disruption of skeletal, craniofacial and cardiac development. Here we review Ryk structure and function focusing on its activity as an axon guidance receptor. (c) 2006 Elsevier Ltd. All rights reserved.

Delayed administration of darbepoetin or erythropoietin protects against ischemic acute renal injury and failure

Administration of human recombinant erythropoietin ( EPO) at time of acute ischemic renal injury ( IRI) inhibits apoptosis, enhances tubular epithelial regeneration, and promotes renal functional recovery. The present study aimed to determine whether darbepoetin-alfa ( DPO) exhibits comparable renoprotection to that afforded by EPO, whether pro or antiapoptotic Bcl-2 proteins are involved, and whether delayed administration of EPO or DPO 6 h following IRI ameliorates renal dysfunction. The model of IRI involved bilateral renal artery occlusion for 45 min in rats ( N = 4 per group), followed by reperfusion for 1-7 days. Controls were sham-operated. Rats were treated at time of ischemia or sham operation ( T0), or post-treated ( 6 h after the onset of reperfusion, T6) with EPO ( 5000 IU/kg), DPO ( 25 mu g/kg), or appropriate vehicle by intraperitoneal injection. Renal function, structure, and immunohistochemistry for Bcl-2, Bcl-XL, and Bax were analyzed. DPO or EPO at T0 significantly abrogated renal dysfunction in IRI animals ( serum creatinine for IRI 0.17 +/- 0.05mmol/l vs DPO-IRI 0.08 +/- 0.03mmol/l vs EPO-IRI 0.04 +/- 0.01mmol/l, P = 0.01). Delayed administration of DPO or EPO ( T6) also significantly abrogated subsequent renal dysfunction ( serum creatinine for IRI 0.17 +/- 0.05mmol/l vs DPO-IRI 0.06 +/- 0.01mmol/l vs EPO-IRI 0.03 +/- 0.03mmol/l, P = 0.01). There was also significantly decreased tissue injury ( apoptosis, P < 0.05), decreased proapoptotic Bax, and increased regenerative capacity, especially in the outer stripe of the outer medulla, with DPO or EPO at T0 or T6. These results reaffirm the potential clinical application of DPO and EPO as novel renoprotective agents for patients at risk of ischemic acute renal failure or after having sustained an ischemic renal insult.

Dietary supplementation of vitamin E and -lipoic acid upregulates cell growth and signaling genes in rat myocardium

The efficacy of antioxidant supplementation in the prevention of cardiovascular disease appears equivocal, however the use of more potent antioxidant combinations than those traditionally used may exert a more positive effect. We have shown previously that supplementation of vitamin E and α-lipoic acid increases cardiac performance during post-ischemia reperfusion in older rats and increases Bcl-2 levels in endothelial cells. The purpose of this study was to examine the effects of vitamin E and α-lipoic acid supplementation on myocardial gene expression with a view to determine their mechanism of action. Young male rats received either a control (n=7) or vitamin E and α-lipoic acid supplemented diet (n=8) for 14 weeks. RNA from myocardial tissue was then amplified and samples were pooled within groups and competitively hybridized to 5K oligonucleotide rat microarrays. The relative expression of each gene was then compared to the control sample. Animals that received the antioxidant-supplemented diet exhibited upregulation (>1.5×) of 13 genes in the myocardium with 2 genes downregulated.� �Upregulated genes include those involved in cell growth and maintenance (LynB, Csf1r, Akt2, Tp53), cell signaling (LynB, Csf1r) and signal transduction (Pacsin2, Csf1r). Downregulated genes encode thyroid (Thrsp) and F-actin binding proteins (Nexilin).

Expression of apoptotic regulatory molecules in renal cell carcinoma: Elevated expression of Fas ligand

Renal cell carcinoma (RCC) is the most common renal neoplasm. Despite being infiltrated by tumour infiltrating lymphocytes (TIL), these TIL are unable to control tumour growth in vivo, suggesting that the cytotoxic capacity of TIL against RCC is impaired, or that the tumour cells are resistant to killing and therefore escape detection by the immune system. It is postulated that the expression of apoptotic regulatory molecules in RCC favours tumour cell survival. The present study has therefore determined the expression of Fas (APO- 1/CD95), Fas ligand (Fas L) and bcl-2 in these tumours. The expression of Fas, Fas L and bcl-2 mRNA transcripts was determined in RCC, normal kidney and peripheral blood by semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR), following RNA extraction and cDNA synthesis from tissues and cell samples. Transcript levels were measured by densitometry after Southern blot hybridization of PCR products with internal radio-labelled oligonucleotide probes; a densitometry score was assigned to each hybridizing DNA band and expressed as a ratio of the glyceraldehyde-3-phosphate dehydrogenase content. In peripheral blood, the expression of Fas L and bcl-2 transcripts was similar between patients and normal healthy individuals; however, Fas transcript expression was significantly down-regulated in the patients' versus normal peripheral blood (P = 0.026). Most interestingly, significantly up-regulated Fas L expression was observed in RCC compared to normal kidney (P = 0.041). In contrast, bcl-2 transcripts were well represented in normal kidney but markedly decreased in RCC (P = 0.021). The expression of Fas transcripts in normal kidney and RCC was variable. These data demonstrate elevated expression of Fas L transcripts in RCC, but the functional relevance of this remains to be investigated.

PERCEPÇÃO DE SUPORTE FAMILIAR E QUALIDADE DE VIDA: UM ESTUDO COM ADOLESCENTES E SEUS PAIS

A família é o primeiro grupo social ao qual o individuo pertence e exerce importante função no desenvolvimento dos seres humanos. Levando-se em consideração o fato da adolescência ser considerada um período do desenvolvimento, marcada por diversas transformações, tanto do individuo quanto do meio familiar, assim como a importância que a família exerce na constituição desses adolescentes, faz-se necessário pensar na qualidade das relações que são estabelecidas. Diante disso, a presente pesquisa teve como objetivo identificar possíveis relações entre a percepção de suporte familiar e qualidade de vida relacionada à saúde de adolescentes e seus responsáveis. Para tanto, buscou-se identificar a percepção dos adolescentes sobre sua qualidade de vida e suporte familiar, bem como a percepção de seus responsáveis sobre os mesmos. Trata-se de um estudo descritivo, quantitativo e de delineamento transversal. Participaram desse estudo 348 sujeitos, sendo 174 adolescentes e 174 responsáveis, escolhidos por conveniência. Dentre os resultados pode-se salientar que os adolescentes possuem percepção de melhor qualidade de vida relacionada à saúde do que seus responsáveis, porém ambos revelam que possuem visão positiva acerca da adolescência. No que se refere à percepção de suporte familiar os dados revelaram que os homens (independente de serem responsáveis ou adolescentes) apresentam melhor percepção acerca da expressão afetiva entre os membros da família e os responsáveis possuem melhor percepção acerca do fator autonomia do que os adolescentes. Conclui se que os objetivos do presente estudo foram atingidos na medida em que foram identificadas inúmeras correlações entre os fatores de percepção de suporte familiar e as dimensões de qualidade de vida relacionada à saúde. Os resultados obtidos sugerem a necessidade de ações que favoreçam o dialogo entre responsáveis e adolescentes favorecendo a aproximação e, consequentemente favorecendo a promoção da saúde.

A novel requirement for C. elegans Alix/ALX-1 in RME-1-mediated membrane transport

BACKGROUND: Alix/Bro1p family proteins have recently been identified as important components of multivesicular endosomes (MVEs) and are involved in the sorting of endocytosed integral membrane proteins, interacting with components of the ESCRT complex, the unconventional phospholipid LBPA, and other known endocytosis regulators. During infection, Alix can be co-opted by enveloped retroviruses, including HIV, providing an important function during virus budding from the plasma membrane. In addition, Alix is associated with the actin cytoskeleton and might regulate cytoskeletal dynamics. RESULTS: Here we demonstrate a novel physical interaction between the only apparent Alix/Bro1p family protein in C. elegans, ALX-1, and a key regulator of receptor recycling from endosomes to the plasma membrane, called RME-1. The analysis of alx-1 mutants indicates that ALX-1 is required for the endocytic recycling of specific basolateral cargo in the C. elegans intestine, a pathway previously defined by the analysis of rme-1 mutants. The expression of truncated human Alix in HeLa cells disrupts the recycling of major histocompatibility complex class I, a known Ehd1/RME-1-dependent transport step, suggesting the phylogenetic conservation of this function. We show that the interaction of ALX-1 with RME-1 in C. elegans, mediated by RME-1/YPSL and ALX-1/NPF motifs, is required for this recycling process. In the C. elegans intestine, ALX-1 localizes to both recycling endosomes and MVEs, but the ALX-1/RME-1 interaction appears to be dispensable for ALX-1 function in MVEs and/or late endosomes. CONCLUSIONS: This work provides the first demonstration of a requirement for an Alix/Bro1p family member in the endocytic recycling pathway in association with the recycling regulator RME-1.

Apoptotic cell‐associated ICAM‐3 in the phagocytic removal of apoptotic cells

Apoptosis is a highly controlled cell death programme that culminates in the exposure of molecular ‘flags’ at the dying cell surface that permit recognition and removal by viable phagocytes. Failure to efficiently remove dying cells can lead to devastating inflammatory and autoimmune disorders. The molecular mechanisms underlying apoptotic cell surface changes are poorly understood. Our previous work has shown an apoptosis-associated functional change in ICAM-3 (a heavily glycosylated, leukocyte-restricted Immunoglobulin Super-Family member) resulting in a molecular ‘flag’ to mediate corpse removal. Here we detail apoptosis-associated changes in ICAM-3 and define their role in ICAM-3’s novel function in apoptotic cell clearance. We show ICAM-3 functions to tether apoptotic leukocytes to macrophages via an undefined receptor. Though CD14 has been suggested as a possible receptor for apoptotic cell-associated ICAM-3, we demonstrate ICAM-3 functions for apoptotic cell clearance in the absence of CD14. Furthermore, we demonstrate leukocytes display early changes in cell surface glycosylation and a marked reduction in ICAM-3, a change that correlates reduced cell volume throughout apoptosis. This loss of ICAM-3 occurs via shedding of ICAM-3 in microparticles (‘apoptotic bodies’). Such microparticles are potent chemoattractants for macrophages. Notably, microparticles from ICAM-3-deficient leukocytes are significantly less chemoattractive than microparticles from their ICAM-3-replete counterparts. These data support the hypothesis that ICAM-3 acts as an apoptotic cell-associated ligand to tether dying cells to phagocytes in a CD14-independent manner. Furthermore our data suggest that released ICAM-3 may promote the recruitment of phagocytes to sites of apoptosis.

Apoptotic cell-derived ICAM-3 promotes both macrophage chemoattraction to and tethering of apoptotic cells

Damaged, aged or unwanted cells are removed from the body by an active process known as apoptosis. This highly orchestrated programme results in cell disassembly and the exposure of ‘flags’ at the dying cell surface that permit recognition and removal by viable cells (phagocytes). Efficient phagocytic removal of dying cells is essential to prevent inflammatory and autoimmune disorders. Relatively little is known of the molecular mechanisms underlying changes at the apoptotic cell surface. We have previously shown that ICAM-3 (a heavily glycosylated, leukocyte-restricted Immunoglobulin Super-Family member) undergoes a change of function as cells die so that it acts as a molecular ‘flag’ to mediate corpse removal. Our work seeks to characterise apoptosis-associated changes in ICAM-3 and define their role in ICAM-3’s novel function in apoptotic cell clearance. Here we extend earlier studies to show that apoptotic cell-associated ICAM-3 functions, at least minimally, to tether apoptotic leukocytes to macrophages via an undefined receptor. Whilst CD14 has been suggested as a possible innate immune receptor for apoptotic cell-associated ICAM-3, we demonstrate ICAM-3 functions for apoptotic cell clearance in the absence of CD14. Our data additionally indicate, that during apoptosis, leukocytes display early changes in cell surface glycosylation and a marked reduction in ICAM-3, a change that correlates with a reduction in cell volume. This reduction in ICAM-3 is explained by cell surface shedding of microparticles (‘apoptotic bodies’) that contain ICAM-3. Such microparticles, released from apoptotic leukocytes, are strongly chemoattractive for macrophages. In addition, microparticles from ICAM-3-deficient leukocytes are significantly less chemoattractive than microparticles from their ICAM-3-replete counterparts. Taken together these data support the hypothesis that ICAM-3 acts as an apoptotic cell-associated ligand to tether dying cells to phagocytes in a CD14-independent manner. Furthermore our data suggest that released ICAM-3 may promote the recruitment of phagocytes to sites of leukocyte apoptosis.

Apoptotic cell-derived ICAM-3 promotes both macrophage chemoattraction to and tethering of apoptotic cells

Damaged, aged or unwanted cells are removed from the body by an active process known as apoptosis. This highly orchestrated programme results in cell disassembly and the exposure of ‘flags’ at the dying cell surface that permit recognition and removal by viable cells (phagocytes). Efficient phagocytic removal of dying cells is essential to prevent inflammatory and autoimmune disorders. Relatively little is known of the molecular mechanisms underlying changes at the apoptotic cell surface. We have previously shown that ICAM-3 (a heavily glycosylated, leukocyte-restricted Immunoglobulin Super-Family member) undergoes a change of function as cells die so that it acts as a molecular ‘flag’ to mediate corpse removal. Our work seeks to characterise apoptosis-associated changes in ICAM-3 and define their role in ICAM-3’s novel function in apoptotic cell clearance. Here we extend earlier studies to show that apoptotic cell-associated ICAM-3 functions, at least minimally, to tether apoptotic leukocytes to macrophages via an undefined receptor. Whilst CD14 has been suggested as a possible innate immune receptor for apoptotic cell-associated ICAM-3, we demonstrate ICAM-3 functions for apoptotic cell clearance in the absence of CD14. Our data additionally indicate, that during apoptosis, leukocytes display early changes in cell surface glycosylation and a marked reduction in ICAM-3, a change that correlates with a reduction in cell volume. This reduction in ICAM-3 is explained by cell surface shedding of microparticles (‘apoptotic bodies’) that contain ICAM-3. Such microparticles, released from apoptotic leukocytes, are strongly chemoattractive for macrophages. In addition, microparticles from ICAM-3-deficient leukocytes are significantly less chemoattractive than microparticles from their ICAM-3-replete counterparts. Taken together these data support the hypothesis that ICAM-3 acts as an apoptotic cell-associated ligand to tether dying cells to phagocytes in a CD14-independent manner. Furthermore our data suggest that released ICAM-3 may promote the recruitment of phagocytes to sites of leukocyte apoptosis.