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Biomedical subjects

R Weil

Publications and source records attributed to R Weil.

At least 19 recordsLinked to original sources

Broadband dielectric and resistivity spectroscopy of WO(3) x H(2)O in the range of 10(3)-10(10) Hz: particle size effect.

The interest in studying the electrical properties of WO(3) x H(2)O powders is made absolutely necessary because their infrared modulation properties depend on their morphologies and electronic populations. Broadband dielectric and resistivity spectra of WO(3) x H(2)O powders were recorded in a frequency range of 10(3)-10(10) Hz at temperatures varying between 200 and 300 K. Complex resistivity and permittivity diagrams have permitted thermal behavior of both dc-conductivity and permittivity to be obtained. A dielectric relaxation is found, attributed to water molecules motions. The role of the powder morphology has been investigated on two types of compounds: the first one being constituted by nanometric particles and the second by micrometric particles. Strong differences are observed in the thermal behaviors of the dc-conductivities (activation energies). Particle size effect is evidenced, giving rise to stronger electron localization on the nanometric particles. The permittivity values and the dynamical behavior of the structural water are also influenced by the particle size effect. A strong interaction between moving polarons and water molecules has been determined.

Journal Article↗

Deciphering the pathway from the TCR to NF-kappaB.

A major regulator of lymphocyte survival and activation is the transcription factor nuclear factor-kappaB (NF-kappaB). Controlled activation of NF-kappaB is essential for the immune and inflammatory response as well as for cell proliferation and protection against apoptosis. The NEMO/IkappaB kinase (IKK) complex is the central integrator of most stimuli leading to NF-kappaB activation, but a detailed knowledge of the upstream events is available only for a limited number of stimuli. In particular, although most players have probably been identified, relatively little is known about the detailed molecular mechanisms involved in the cascade leading to NF-kappaB activation following engagement of the T-cell receptor by a foreign antigen. In this review, we discuss recent insights into this specific signal transduction cascade, and the way it is controlled both spatially and temporally.

Adaptor Proteins, Signal Transducing↗

A follow up study of patients with paraneoplastic neurological disease in the United Kingdom.

OBJECTIVES: To examine the range of clinical phenotypes, tumour associations, relevant investigations, response to therapy and outcome in a large series of non-selected patients with paraneoplastic neurological disease (PND) affecting the central nervous system (CNS) in the United Kingdom. METHODS: Data were obtained on patients either through direct referral or through the British Neurological Surveillance Unit (BNSU) from February 2000 to January 2001. Physicians were asked to supply information about age and sex of patients, presenting neurological syndromes, the basis of the diagnosis of PND, any associated malignancy, and treatment. Case notes were reviewed and follow up data obtained where possible one year after notification. RESULTS: A total of 63 patients (48 females, 15 males) were identified, 48 through the BNSU and 15 through direct referral. Of these 52 were diagnosed as having definite PND, 10 probable PND, and 1 possible PND. The median age of onset of PND was 66 years (range 30-80 years) and only 7 patients (11%) were less than 50 years at presentation. In 53 patients (84%) the PND preceded the diagnosis of cancer. Paraneoplastic sensory neuronopathy, paraneoplastic encephalomyelitis, and paraneoplastic cerebellar degeneration (PCD) were the most common syndromes reported. The benefit of magnetic resonance imaging in the diagnosis of the disease was limited, while fluorodeoxyglucose positron emission tomography was shown to be useful for the detection of an occult malignancy in 10 out of 14 patients. Antineuronal antibodies were positive in 44/57 (77%) of cases. The following tumours were diagnosed: small cell lung cancer (30%), breast cancer (14%), ovarian cancer (8%), non-small cell lung cancer (8%), Hodgkin's lymphoma (6%), other (16%). With the exception of PCD associated with mesothelioma all other tumours diagnosed in these patients had been previously documented as being associated with PND. Only treatment of the tumour was found to be associated with a stable or improved neurological outcome at last follow up (Fisher's exact test = 4.7, p<0.03). Median survival time was 43 months (95% CI 28 to 57) from onset of neurological disease as calculated using the Kaplan-Meier survival analysis. CONCLUSIONS: PND has a striking female preponderance usually affecting patients in their sixth decade and above. The median survival in our study was 43 months. The majority of patients with PND are not known to have cancer at the time of diagnosis. Our study confirms the importance of diagnosing and treating the underlying tumour.

Adult↗

Characterization of an IL-2 mimetic with therapeutic potential.

Human interleukin-2 (IL-2) interacts with two types of functional receptors (IL-2R alpha betagamma and IL-2R betagamma) and acts on a broad range of target cells involved in inflammatory reactions and immune responses. IL-2 is also used in different clinical trials aimed at improving the treatment of some cancers and the recovery of CD4 lymphocytes by HIV patients. The therapeutic index of IL-2 is limited by various side effects dominated by the vascular leak syndrome. We have shown that a chemically synthesised fragment of the IL-2 sequence can fold into a helical tetramer likely mimicking the quatemary structure of an hemopoietin. Indeed, peptide p1-30 (containing amino acids 1 to 30, including the sequence corresponding to the entire alpha helix A of IL-2) spontaneously folds into an alpha-helical homotetramer and stimulates the growth of T-cell lines expressing human IL-2R beta, whereas shorter versions of the peptide lack helical structure and are inactive. At the cellular level, p1-30 induces lymphokine-activated killer (LAK) cells and preferentially activates CD8 low lymphocytes and natural killer cells, which constitutively express IL-2R beta. A significant IFN-gamma production is also detected following p1-30 stimulation. A mutant form of p1-30 (Asp20-->Lys) which is likely unable to induce vascular leak syndrome remains capable to generate LAK cells like the original p1-30 peptide. Altogether our data suggest that p1-30 has therapeutic potential.

CD8-Positive T-Lymphocytes↗

Phorbol esters and cytokines regulate the expression of the NEMO-related protein, a molecule involved in a NF-kappa B-independent pathway.

The NF-kappaB signaling pathway plays a crucial role in the immune, inflammatory, and apoptotic responses. Recently, we identified the NF-kappaB Essential Modulator (NEMO) as an essential component of this pathway. NEMO is a structural and regulatory subunit of the high molecular kinase complex (IKK) responsible for the phosphorylation of NF-kappaB inhibitors. Data base searching led to the isolation of a cDNA encoding a protein we called NRP (NEMO-related protein), which shows a strong homology to NEMO. Here we show that NRP is present in a novel high molecular weight complex, that contains none of the known members of the IKK complex. Consistently, we could not observe any effect of NRP on NF-kappaB signaling. Nonetheless, we could demonstrate that treatment with phorbol esters induces NRP phosphorylation and decreases its half-life. This phosphorylation event could only be inhibited by K-252a and stauroporin. We also show that de novo expression of NRP can be induced by interferon and tumor necrosis factor alpha and that these two stimuli have a synergistic effect on NRP expression. In addition, we observed that endogenous NRP is associated with the Golgi apparatus. Analogous to NEMO, we find that NRP is associated in a complex with two kinases, suggesting that NRP could play a similar role in another signaling pathway.

Amino Acid Sequence↗

The first alpha helix of interleukin (IL)-2 folds as a homotetramer, acts as an agonist of the IL-2 receptor beta chain, and induces lymphokine-activated killer cells.

Interleukin (IL)-2 interacts with two types of functional receptors (IL-2Ralphabetagamma and IL-2Rbetagamma) and acts on a broad range of target cells involved in inflammatory reactions and immune responses. For the first time, we show that a chemically synthesized fragment of the IL-2 sequence can fold into a molecule mimicking the quaternary structure of a hemopoietin. Indeed, peptide p1-30 (containing amino acids 1-30, covering the entire alpha helix A of IL-2) spontaneously folds into an alpha-helical homotetramer and stimulates the growth of T cell lines expressing human IL-2Rbeta, whereas shorter versions of the peptide lack helical structure and are inactive. We also demonstrate that this neocytokine interacts with a previously undescribed dimeric form of IL-2Rbeta. In agreement with its binding to IL-2Rbeta, p1-30 activates Shc and p56(lck) but unlike IL-2, fails to activate Janus kinase (Jak)1, Jak3, and signal transducer and activator of transcription 5 (STAT5). Unexpectedly, we also show that p1-30 activates Tyk2, thus suggesting that IL-2Rbeta may bind to different Jaks depending on its oligomerization. At the cellular level, p1-30 induces lymphokine-activated killer (LAK) cells and preferentially activates CD8(low) lymphocytes and natural killer cells, which constitutively express IL-2Rbeta. A significant interferon gamma production is also detected after p1-30 stimulation. A mutant form of p1-30 (Asp20-->Lys), which is likely unable to induce vascular leak syndrome, remains capable of generating LAK cells, like the original p1-30 peptide. Altogether, our data suggest that p1-30 has therapeutic potential.

Amino Acid Sequence↗

PKR stimulates NF-kappaB irrespective of its kinase function by interacting with the IkappaB kinase complex.

The interferon (IFN)-induced double-stranded RNA-activated protein kinase PKR mediates inhibition of protein synthesis through phosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eIF2alpha) and is also involved in the induction of the IFN gene through the activation of the transcription factor NF-kappaB. NF-kappaB is retained in the cytoplasm through binding to its inhibitor IkappaBalpha. The critical step in NF-kappaB activation is the phosphorylation of IkappaBalpha by the IkappaB kinase (IKK) complex. This activity releases NF-kappaB from IkappaBalpha and allows its translocation to the nucleus. Here, we have studied the ability of PKR to activate NF-kappaB in a reporter assay and have shown for the first time that two catalytically inactive PKR mutants, PKR/KR296 and a deletion mutant (PKR/Del42) which lacks the potential eIF2alpha-binding domain, can also activate NF-kappaB. This result indicated that NF-kappaB activation by PKR does not require its kinase activity and that it is independent of the PKR-eIF2alpha relationship. Transfection of either wild-type PKR or catalytically inactive PKR in PKR(0/0) mouse embryo fibroblasts resulted in the activation of the IKK complex. By using a glutathione S-transferase pull-down assay, we showed that PKR interacts with the IKKbeta subunit of the IKK complex. This interaction apparently does not require the integrity of the IKK complex, as it was found to occur with extracts from cells deficient in the NF-kappaB essential modulator, one of the components of the IKK complex. Therefore, our results reveal a novel pathway by which PKR can modulate the NF-kappaB signaling pathway without using its kinase activity.

Amino Acid Sequence↗

Well-done, grilled red meat increases the risk of colorectal adenomas.

Red meat or meat-cooking methods such as frying and doneness level have been associated with an increased risk of colorectal and other cancers. It is unclear whether it is red meat intake or the way it is cooked that is involved in the etiology of colorectal cancer. To address this issue, we developed an extensive food frequency questionnaire module that collects information on meat-cooking techniques as well as the level of doneness for individual meat items and used it in a study of colorectal adenomas, known precursors of colorectal cancer. A case-control study of colorectal adenomas was conducted at the National Naval Medical Center (Bethesda, MD) between April 1994 and September 1996. All cases (n = 146) were diagnosed with colorectal adenomas at sigmoidoscopy or colonoscopy and histologically confirmed. Controls (n = 228) were screened with sigmoidoscopy and found not to have colorectal adenomas. The subjects completed a food frequency questionnaire and answered detailed questions on meat-cooking practices. We used frequency and portion size to estimate grams of meat consumed per day for total meat as well as for meat subgroups defined by cooking methods and doneness levels. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using logistic regression, adjusted for age, gender, total caloric intake, reason for screening (routine or other), and several established risk factors for colorectal adenomas or cancer, including the use of nonsteroidal anti-inflammatory drugs, physical activity, and pack-years of cigarette smoking. There was an increased risk of 11% per 10 g/day (or 2.5 oz/week) of reported red meat consumption (OR, 1.11; CI, 1.03-1.19). The increased risk was mainly associated with well-done/very well-done red meat, with an excess risk of 29% per 10 g/day (OR, 1.29; CI, 1.08-1.54) versus an excess of 10% per 10 g/day (OR, 1.10; CI, 0.96-1.26) for consumption of rare/medium red meat. High-temperature cooking methods were also associated with increased risk; 26% per 10 g/day (OR, 1.26; CI, 1.06-1.50) of grilled red meat and 15% per 10 g/day (OR, 1.15; CI, 0.97-1.36) of pan-fried red meat consumption. There was an increased risk of colorectal adenomas associated with higher intake of red meat, most of which was due to the subgroup of red meat that was cooked until well done/very well done and/or by high-temperature cooking techniques, such as grilling. These results are consistent with the hypothesis that carcinogenic compounds formed by high-temperature cooking techniques, such as heterocyclic amines and polycyclic aromatic hydrocarbons, may contribute to the risk of developing colorectal tumors.

Adenoma↗

Morphologic and genetic analysis of retinal angioma associated with massive gliosis in a patient with von Hippel-Lindau disease.

We report morphologic and genetic analysis of bilateral retinal angiomas in a 35-year-old patient with von Hippel-Lindau (VHL) disease. Enucleation of both eyes revealed extensive intraocular tumor. Whereas the right eye demonstrated large amounts of retinal angioma tissue, the left eye showed small areas of retinal angioma associated with massive diffuse retinal gliosis. Genetic analysis of the angioma showed allelic deletion of the VHL gene locus, suggesting that the origin of the angiomas was directly related to the patient's underlying VHL disease. Genetic analysis of the pleomorphic glial proliferation showed no allelic VHL gene deletion, which is consistent with the assessment that the glial component represents a reactive process. Apoptosis detected by TUNEL revealed lack of DNA fragmentation in the angioma; in contrast, many positive signals were found in the massive gliosis. We confirmed that the abnormal VHL genes were located in the "stromal" cells of the retinal angioma. Massive gliosis in VHL disease is a true reactive retinal gliosis.

Adult↗

Altered expression of tyrosine kinases of the Src and Syk families in human T-cell leukemia virus type 1-infected T-cell lines.

During the late phase of adult T-cell leukemia/lymphoma, a severe lymphoproliferative disorder caused by human T-cell leukemia virus type 1 (HTLV-1), leukemic cells no longer produce interleukin-2. Several studies have reported the lack of the Src-like protein tyrosine kinase Lck and overexpression of Lyn and Fyn in these cells. In this report we demonstrate that, in addition to the downregulation of TCR, CD45, and Lck (which are key components of T-cell activation), HTLV-1-infected cell lines demonstrate a large increase of FynB, a Fyn isoform usually poorly expressed in T cells. Furthermore, similar to anergic T cells, Fyn is hyperactive in one of these HTLV-1-infected T-cell lines, probably as a consequence of Csk downregulation. A second family of two proteins, Zap-70 and Syk, relay the signal of T-cell activation. We demonstrate that in contrast to uninfected T cells, Zap-70 is absent in HTLV-1-infected T cells, whereas Syk is overexpressed. In searching for the mechanism responsible for FynB overexpression and Zap-70 downregulation, we have investigated the ability of the Tax and Rex proteins to modulate Zap-70 expression and the alternative splicing mechanism which gives rise to either FynB or FynT. By using Jurkat T cells stably transfected with the tax and rex genes or inducibly expressing the tax gene, we found that the expression of Rex was necessary to increase fynB expression, suggesting that Rex controls fyn gene splicing. Conversely, with the same Jurkat clones, we found that the expression of Tax but not Rex could downregulate Zap-70 expression. These results suggest that the effect of Tax and Rex must cooperate to deregulate the pathway of T-cell activation in HTLV-1-infected T cells.

Down-Regulation↗

Direct association and nuclear import of the hepatitis B virus X protein with the NF-kappaB inhibitor IkappaBalpha.

The X protein of hepatitis B virus (HBV) is a transcriptional activator which is required for infection and may play an important role in HBV-associated hepatocarcinogenesis. It has been suggested that X acts as a nuclear coactivator or stimulates several signal transduction pathways by acting in the cytoplasm. One of these pathways leads to the nuclear translocation of NF-kappaB. A recent report indicates that X activates NF-kappaB by acting on two cytoplasmic inhibitors of this family of transcription factors: IkappaBalpha and the precursor/inhibitor p105. We demonstrate here that X directly interacts with IkappaBalpha, which is able to transport it to the nucleus by a piggyback mechanism. This transport requires a region of IkappaBalpha (the second ankyrin repeat) which has been demonstrated to be involved in its nuclear import following NF-kappaB activation. Using deletion mutants, we showed that amino acids 249 to 253 of IkappaBalpha (located in the C-terminal part of the sixth ankyrin repeat) play a critical role in the interaction with X. This small region overlaps one of the domains of IkappaBalpha mediating the interaction with the p50 and p65 subunits of NF-kappaB and is also close to the nuclear export sequence of IkappaBalpha, therefore providing a potential explanation for the nuclear accumulation of IkappaBalpha with X. This association can also be observed upon the induction of endogenous IkappaBalpha by tumor necrosis factor alpha (TNF-alpha) treatment of Chang cells expressing X. In accordance with this observation, band shift analysis indicates that X induces a sustained NF-kappaB activation following TNF-alpha treatment, probably by preventing the reassociation of newly synthesized nuclear IkappaBalpha with DNA-bound NF-kappaB complexes.

Animals↗

Multiple endocrine neoplasia type 1: atypical presentation, clinical course, and genetic analysis of multiple tumors.

Multiple endocrine neoplasia type 1 (MEN1) is characterized by the development of endocrine tumors of the parathyroid and pituitary glands, pancreas, and duodenum. Less frequently occurring tumors associated with MEN1 include non-endocrine tumors such as lipomas and angiofibromas. An increased incidence of thyroid neoplasms, leiomyomas, adrenal cortical hyperplasia, hepatic focal nodular hyperplasia, and renal angiomyolipoma has been noted in the MEN1 population. The pathogenesis of non-neuroendocrine tumors in MEN1 is unknown. We report a complex clinical course and a detailed morphologic and genetic analysis of a series of tumors that developed in a patient with MEN1. All tumors were microdissected and analyzed for loss of heterozygosity of the MEN1 gene. A germline mutation of the MEN1 gene was detected, and deletions of the MEN1 gene were consistently detected in multiple neuroendocrine tumors involving the parathyroid glands and the pancreas and a hepatic neuroendocrine tumor metastasis, as predicted by Knudson's "two hit" hypothesis. Two hits of the MEN1 gene were also detected in esophageal leiomyoma tissue, suggesting that tumorigenesis was directly related to the patient's underlying MEN1. In contrast, follicular thyroid adenoma, papillary thyroid carcinoma, hepatic focal nodular hyperplasia, and adrenal cortical hyperplasia consistently showed retained heterozygosity of the MEN1 gene with flanking markers and an intragenic marker. Therefore, these tumors appear to develop along pathogenetic pathways that are different from classical MEN1-associated tumors.

Adult↗

Complementation cloning of NEMO, a component of the IkappaB kinase complex essential for NF-kappaB activation.

We have characterized a flat cellular variant of HTLV-1 Tax-transformed rat fibroblasts, 5R, which is unresponsive to all tested NF-kappaB activating stimuli, and we report here its genetic complementation. The recovered full-length cDNA encodes a 48 kDa protein, NEMO (NF-kappaB Essential MOdulator), which contains a putative leucine zipper motif. This protein is absent from 5R cells, is part of the high molecular weight IkappaB kinase complex, and is required for its formation. In vitro, NEMO can homodimerize and directly interacts with IKK-2. The NEMO cDNA was also able to complement another NF-kappaB-unresponsive cell line, 1.3E2, in which the protein is also absent, allowing us to demonstrate that this factor is required not only for Tax but also for LPS, PMA, and IL-1 stimulation of NF-kappaB activity.

Amino Acid Sequence↗

Cytosol is the prime compartment of hepatitis B virus X protein where it colocalizes with the proteasome.

The hepatitis B virus X protein plays an important role in the regulation of viral genome expression and has also been implicated in the development of liver cancer associated with chronic viral infection. Several effects have been attributed to X but their biological relevance remains elusive. One of the confusing issues has been so far the uncertainty concerning its cellular location. To gain insight into the mechanism(s) how X exerts its effects, we have analysed its subcellular distribution and its dependency on the cell cycle. We used two complementary approaches namely, immunolocalization using a cell line stably expressing X, and characterization of the dynamics of X location in living cells by means of the reporter gene GFP. Our data clearly define the cytosol as the prime location of X, irrespectively of the cell cycle and show in addition the close attachment of a fraction of X to the nuclear membrane. However, X does not associate with any cytoplasmic vesicles and organelles so far tested. In contrast, our study provides strong evidence for the codistribution of X with the cytosolic fraction of proteasomes. In pulse-chase experiments, X decayed with a half-life of less than 30 min and proteasome-inhibitors did not modify its turnover, suggesting that X colocalization with the proteasome does not simply point to its degradation pathway. The proteolytic processing of the p105 precursor of the p50 subunit of the NF-kappaB transcription factor, which has been shown to be proteasome-dependent, is markedly slow down in the presence of X. These findings suggest that X modulates the processing rate of p105 by acting presumably at the level of the proteasome. Thus, targeting of proteasomes by X might be one of the pathways employed by this viral protein to subvert cellular functions.

Animals↗

Mutations of the MEN1 tumor suppressor gene in pituitary tumors.

Although pituitary adenomas are monoclonal proliferations, somatic mutations involving genes that govern cell proliferation or hormone production have been difficult to identify. The genetic etiology of most pituitary tumors, therefore, remains unknown. Pituitary adenomas can develop sporadically or as a part of multiple endocrine neoplasia type 1 (MEN1). Recently, the gene responsible for MEN1 was cloned. To elucidate the potential etiological role of the MEN1 gene in pituitary tumorigenesis, 39 sporadic pituitary adenomas from 38 patients and 1 pituitary adenoma from a familial MEN1 patient were examined for MEN1 gene mutations and allelic deletions. Four of 39 sporadic pituitary adenomas showed a deletion of one copy of the MEN1 gene, and a specific MEN1 gene mutation in the remaining gene copy was detected in 2 of these tumors. The corresponding germ-line sequence was normal in all sporadic cases. A specific MEN1 mutation was detected in a pituitary adenoma and corresponding germ-line DNA in a patient with familial MEN1. An allelic deletion of the remaining copy of the MEN1 gene was also found in the patient's tumor. Genetic alterations of the MEN1 gene represent a candidate pathogenetic mechanism of pituitary tumorigenesis. The data suggest that somatic MEN1 gene mutations and deletions play a causative role in the development of a subgroup of sporadic pituitary adenomas.

Adenoma↗

Regulation of IkappaBbeta degradation. Similarities to and differences from IkappaBalpha.

The transcription factor NF-kappaB (nuclear factor-kappaB) is neutralized in nonstimulated cells through cytoplasmic retention by IkappaB inhibitors. In mammalian cells, two major forms of IkappaB proteins, IkappaBalpha and IkappaBbeta, have been identified. Upon treatment with a large variety of inducers, IkappaBalpha and IkappaBbeta are proteolytically degraded, resulting in NF-kappaB translocation into the nucleus. Recent observations suggest that phosphorylation of serines 32 and 36 and subsequent ubiquitination of lysines 21 and 22 of IkappaBalpha control its signal-induced degradation. In this study we provide evidence that critical residues in the NH2-terminal region of IkappaBbeta (serines 19 and 23) as well as its COOH-terminal PEST region control IkappaBbeta proteolysis. However Lys-9, the unique lysine residue in the NH2-terminal region of IkappaBbeta, is not absolutely required for its degradation. We also demonstrate that following stimulation, an underphosphorylated nondegradable form of IkappaBbeta accumulates. Surprisingly, our data suggest that unlike IkappaBalpha, IkappaBbeta is constitutively phosphorylated on one or two of the critical NH2-terminal serine residues. Thus, phosphorylation of these sites is necessary for degradation but does not necessarily constitute the signal-induced event that targets the molecule for proteolysis.

Animals↗