Remarkable interest has risen in the idea that oxidative/nitrosative stress is mediated in the etiology of numerous human diseases. Oxidative/Nitrosative stress is the result of an disequilibrium in oxidant/antioxidant which reveals from continuous increase of Reactive Oxygen and Reactive Nitrogen Species production. The aim of this review is to emphasize with current information the importance of antioxidants which play the role in cellular responce against oxidative/nitrosative stress, which would be helpful in enhancing the knowledge of any biochemist, pathophysiologist, or medical personnel regarding this important issue. Products of lipid peroxidation have commonly been used as biomarkers of oxidative/nitrosative stress damage. Lipid peroxidation generates a variety of relatively stable decomposition end products, mainly α, β-unsaturated reactive aldehydes, such as malondialdehyde, 4-hydroxy-2-nonenal, 2-propenal (acrolein) and isoprostanes, which can be measured in plasma and urine as an indirect index of oxidative/nitrosative stress. Antioxidants are exogenous or endogenous molecules that mitigate any form of oxidative/nitrosative stress or its consequences. They may act from directly scavenging free radicals to increasing antioxidative defences. Antioxidant deficiencies can develop as a result of decreased antioxidant intake, synthesis of endogenous enzymes or increased antioxidant utilization. Antioxidant supplementation has become an increasingly popular practice to maintain optimal body function. However, antoxidants exhibit pro-oxidant activity depending on the specific set of conditions. Of particular importance are their dosage and redox conditions in the cell.

The stimulation of insulin secretion from the beta cells of the islets of Langerhans appears to be mediated by a decrease in the cell-membrane potassium-ion permeability. Tolbutamide reduced K+ movement through an ATP-sensitive K+ channel in patches of plasma membrane from an insulin-producing cell line when applied to the external surface of the membrane. The effect occurred at concentrations which exist in the serum of patients treated with tolbutamide and which stimulate insulin secretion from islets of Langerhans in vitro. Glibenclamide had a similar effect but, in keeping with its greater therapeutic potency, at concentrations one hundred times lower. These findings suggest that an ATP-sensitive K+ channel or a protein closely associated with it may be the receptor through which sulphonylureas act to stimulate insulin secretion in vitro.

The generation of B-lymphocytes from hematopoietic stem cells is controlled by multiple transcription factors regulating distinct developmental aspects. Ikaros and PU.1 act in parallel pathways to control the development of lymphoid progenitors in part by regulating the expression of essential signaling receptors (Flt3, c-Kit, and IL-7R alpha). The generation of the earliest B cell progenitors depends on E2A and EBF, which coordinately activate the B cell gene expression program and immunoglobulin heavy-chain gene rearrangements at the onset of B-lymphopoiesis. Pax5 restricts the developmental options of lymphoid progenitors to the B cell lineage by repressing the transcription of lineage-inappropriate genes and simultaneously activating the expression of B-lymphoid signaling molecules. LEF1 and Sox4 contribute to the survival and proliferation of pro-B cells in response to extracellular signals. Finally, IRF4 and IRF8 together control the termination of pre-B cell receptor signaling and thus promote differentiation to small pre-B cells undergoing light-chain gene rearrangements.

NF-kappa B is a widely used regulator of inducible and tissue-specific gene control. In the cytosol, when complexed to an inhibitory molecule, I kappa B, NF-kappa B is in an inactive form and cannot bind DNA. Activation of cells with appropriate stimuli results in the dissociation of NF-kappa B from I kappa B and its translocation to the nucleus as an active binding protein. We now demonstrate that NF-kappa B binding in vitro can be inhibited by agents that modify free sulfhydryls. Binding is eliminated after treatment with N-ethylmaleimide, an alkylating agent, and diamide, an oxidizing agent. The diamide effect can be reversed by 2-mercaptoethanol. Further, 2-mercaptoethanol acts synergistically with deoxycholate plus Nonidet P-40 in converting inactive cytosolic NF-kappa B to an active DNA-binding form. It is therefore possible that modulation of the redox state of NF-kappa B could represent a post-translational control mechanism for this factor.

The Raf kinase inhibitor protein (RKIP) acts as a negative regulator of the mitogen-activated protein (MAP) kinase (MAPK) cascade initiated by Raf-1. RKIP inhibits the phosphorylation of MAP/extracellular signal-regulated kinase 1 (MEK1) by Raf-1 by disrupting the interaction between these two kinases. We show here that RKIP also antagonizes the signal transduction pathways that mediate the activation of the transcription factor nuclear factor kappa B (NF-kappaB) in response to stimulation with tumor necrosis factor alpha (TNF-alpha) or interleukin 1 beta. Modulation of RKIP expression levels affected NF-kappaB signaling independent of the MAPK pathway. Genetic epistasis analysis involving the ectopic expression of kinases acting in the NF-kappaB pathway indicated that RKIP acts upstream of the kinase complex that mediates the phosphorylation and inactivation of the inhibitor of NF-kappaB (IkappaB). In vitro kinase assays showed that RKIP antagonizes the activation of the IkappaB kinase (IKK) activity elicited by TNF-alpha. RKIP physically interacted with four kinases of the NF-kappaB activation pathway, NF-kappaB-inducing kinase, transforming growth factor beta-activated kinase 1, IKKalpha, and IKKbeta. This mode of action bears striking similarities to the interactions of RKIP with Raf-1 and MEK1 in the MAPK pathway. Emerging data from diverse organisms suggest that RKIP and RKIP-related proteins represent a new and evolutionarily highly conserved family of protein kinase regulators. Since the MAPK and NF-kappaB pathways have physiologically distinct roles, the function of RKIP may be, in part, to coordinate the regulation of these pathways.

Bacteriophage T4 has a very efficient mechanism for infecting cells. The key component of this process is the baseplate, located at the end of the phage tail, which regulates the interaction of the tail fibres and the DNA ejection machine. A complex of gene product (gp) 5 (63K) and gp27 (44K), the central part of the baseplate, is required to penetrate the outer cell membrane of Escherichia coli and to disrupt the intermembrane peptidoglycan layer, promoting subsequent entry of phage DNA into the host. We present here a crystal structure of the (gp5-gp27)3 321K complex, determined to 2.9 A resolution and fitted into a cryo-electron microscopy map at 17 A resolution of the baseplate-tail tube assembly. The carboxy-terminal domain of gp5 is a triple-stranded beta-helix that forms an equilateral triangular prism, which acts as a membrane-puncturing needle. The middle lysozyme domain of gp5, situated on the periphery of the prism, serves to digest the peptidoglycan layer. The amino-terminal, antiparallel beta-barrel domain of gp5 is inserted into a cylinder formed by three gp27 monomers, which may serve as a channel for DNA ejection.

Mammalian target of rapamycin (mTOR) is a key regulator of cell growth that associates with raptor and rictor to form the mTOR complex 1 (mTORC1) and mTORC2, respectively. Raptor is required for oxidative muscle integrity, whereas rictor is dispensable. In this study, we show that muscle-specific inactivation of mTOR leads to severe myopathy, resulting in premature death. mTOR-deficient muscles display metabolic changes similar to those observed in muscles lacking raptor, including impaired oxidative metabolism, altered mitochondrial regulation, and glycogen accumulation associated with protein kinase B/Akt hyperactivation. In addition, mTOR-deficient muscles exhibit increased basal glucose uptake, whereas whole body glucose homeostasis is essentially maintained. Importantly, loss of mTOR exacerbates the myopathic features in both slow oxidative and fast glycolytic muscles. Moreover, mTOR but not raptor and rictor deficiency leads to reduced muscle dystrophin content. We provide evidence that mTOR controls dystrophin transcription in a cell-autonomous, rapamycin-resistant, and kinase-independent manner. Collectively, our results demonstrate that mTOR acts mainly via mTORC1, whereas regulation of dystrophin is raptor and rictor independent.

Wnt glycoproteins regulate homeostasis and development by binding to membrane Frizzled-LRP5/6 receptor complexes. Wnt signaling includes a canonical pathway involving cytosolic beta-catenin stabilization, nuclear translocation and gene regulation, acting as a co-activator of T-cell factor (TCF) proteins, and noncanonical pathways that activate Rho, Rac, JNK and PKC, or modulate Ca(2+) levels. DICKKOPF-1 (DKK-1) encodes a secreted Wnt antagonist that binds to LRP5/6 and induces its endocytosis, leading to inhibition of the canonical pathway. We show that activation of canonical signaling by Wnt1 or ectopic expression of active beta-catenin, TCF4 or LRP6 mutants induces transcription of the human DKK-1 gene. Multiple beta-catenin/TCF4 sites in the DKK-1 gene promoter contribute to this activation. In contrast, Wnt5a, which signals through noncanonical pathways, does not activate DKK-1. Northern and Western blot studies show that activation of the Wnt/beta-catenin pathway by treatment with lithium or Wnt3a-conditioned medium, or by stable expression of either Wnt1 or beta-catenin, increases DKK-1 RNA and protein, thus initiating a negative feedback loop. However, we found that DKK-1 expression decreases in human colon tumors, which suggests that DKK-1 acts as a tumor suppressor gene in this neoplasia. Our data indicate that the Wnt/beta-catenin pathway is downregulated by the induction of DKK-1 expression, a mechanism that is lost in colon cancer.

Interleukin (IL) 6 is a proinflammtory cytokine produced by antigen-presenting cells and nonhematopoietic cells in response to external stimuli. It was initially identified as a B cell growth factor and inducer of plasma cell differentiation in vitro and plays an important role in antibody production and class switching in vivo. However, it is not clear whether IL-6 directly affects B cells or acts through other mechanisms. We show that IL-6 is sufficient and necessary to induce IL-21 production by naive and memory CD4(+) T cells upon T cell receptor stimulation. IL-21 production by CD4(+) T cells is required for IL-6 to promote B cell antibody production in vitro. Moreover, administration of IL-6 with inactive influenza virus enhances virus-specific antibody production, and importantly, this effect is dependent on IL-21. Thus, IL-6 promotes antibody production by promoting the B cell helper capabilities of CD4(+) T cells through increased IL-21 production. IL-6 could therefore be a potential coadjuvant to enhance humoral immunity.

The race to produce vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) began when the first sequence was published, and this forms the basis for vaccines currently deployed globally. Independent lineages of SARS-CoV-2 have recently been reported: UK, B.1.1.7; South Africa, B.1.351; and Brazil, P.1. These variants have multiple changes in the immunodominant spike protein that facilitates viral cell entry via the angiotensin-converting enzyme-2 (ACE2) receptor. Mutations in the receptor recognition site on the spike are of great concern for their potential for immune escape. Here, we describe a structure-function analysis of B.1.351 using a large cohort of convalescent and vaccinee serum samples. The receptor-binding domain mutations provide tighter ACE2 binding and widespread escape from monoclonal antibody neutralization largely driven by E484K, although K417N and N501Y act together against some important antibody classes. In a number of cases, it would appear that convalescent and some vaccine serum offers limited protection against this variant.

Keywords: ACE2; B.1.351; SARS-CoV-2; South Africa; antibody; escape; neutralization; receptor-binding domain; vaccine; variant.

The posttranscriptional regulator (p27x-III) of human T cell leukemia virus type I (HTLV-I) is located predominantly in the cell nucleolus. A highly basic amino-terminal sequence (NH2-Met-Pro-Lys-Thr-Arg-Arg-Arg-Pro-Arg-Arg-Ser-Gln-Arg-Lys-Arg-Pro-Pro -Thr- Pro) in this protein, when fused to the amino termini of beta-galactosidase and p40x of HTLV-I, acts as an autonomous signal capable of directing the hybrid proteins to the cell nucleolus.

It is currently believed that latently infected, resting B lymphocytes are central to gammaherpesvirus persistence, whereas mucosal epithelial cells are considered nonessential. We have readdressed the question of nonlymphoid persistence using murine gammaherpesvirus 68 (MHV-68). To dissect lymphoid from nonlymphoid persistence, we used microMT transgenic mice that are defective in B cells. MHV-68 DNA persisted in the lungs of intact and B cell-deficient mice. Both episomal and linear forms of the virus genome were present in lungs, implying the presence of both latency and productive replication. In situ hybridization for virus tRNA transcripts revealed latent MHV-68 in pulmonary epithelial cells. Infectious virus was recovered from the lungs of microMT mice after T cell depletion, showing that the persisting virus DNA was reactivatable. Finally, using adoptive transfer of B cells into B cell-deficient mice, it was shown that virus persisting in lungs seeded splenic B cells, and virus resident in the spleen seeded the lungs. These results show that mucosal epithelia can act as a nonlymphoid reservoir for gammaherpesvirus persistence, and that there is a two-way movement of virus between lymphoid and nonlymphoid compartments during persistence.

The herpes simplex virus (HSV-1) immediate early protein ICP27 is a regulatory protein which is essential for virus replication. The phenotype of temperature-sensitive and deletion mutants in ICP27 includes overexpression of some immediate early and early gene products and greatly reduced levels of late gene products. To determine whether regulation by ICP27 occurs primarily at the transcriptional level, we have studied the expression of two immediate early products (ICP4 and ICP27) and two late gene products (glycoprotein B and glycoprotein C) at the level of transcription initiation, accumulation of steady state mRNA, and protein synthesis in an ICP27 temperature-sensitive mutant tsLG4, compared to wild-type HSV-1. At the nonpermissive temperature in tsLG4-infected cells, the two immediate early gene products ICP4 and ICP27 were overexpressed both at the mRNA and protein level although synthesis of these transcripts as measured by nuclear runoff assays was reduced relative to the wild-type HSV-1 infections. The transcription of late gene products glycoprotein B (gB) and glycoprotein C (gC) was lower in runoff assays from tsLG4 infections suggesting that the reduction in the level of late products occurred at the transcriptional level. However, temperature shift experiments in which tsLG4-infected cells were shifted to the nonpermissive temperature at various times after infection showed that the synthesis of late transcripts was not altered 2 hr after the shift whereas both the accumulation of leaky late and late mRNA and the incorporation of [35S]methionine into newly synthesized gB and gC was reduced by 2 hr after the shift to nonpermissive temperature. Therefore, while the synthesis of new transcripts continued, the accumulation of late mRNAs and their translation into protein was reduced when ICP27 was defective, whereas, the converse was found for immediate early products. That is, the synthesis of new transcripts was reduced yet mRNA and protein accumulated to high levels. These results suggest that ICP27 acts at least in part post-transcriptionally to regulate the expression of immediate early and late gene products.

Increasing evidence has demonstrated striking sex differences in the pathophysiology of and outcome after acute neurological injury. Lesser susceptibility to postischemic and posttraumatic brain injury in females has been observed in experimental models. Additional evidence suggests this sex difference extends to humans as well. The greater neuroprotection afforded to females is likely due to the effects of circulating estrogens and progestins. In fact, exogenous administration of both hormones has been shown to improve outcome after cerebral ischemia and traumatic brain injury in experimental models. The neuroprotection provided by periinjury administration of these hormones extends to males as well. The mechanisms by which estrogen and progesterone provide such neuroprotection are likely multifactorial, and probably depend on the type and severity of injury as well as the type and concentration of hormone present. Both genomic and nongenomic mechanisms may be involved. Estrogen's putative effects include preservation of autoregulatory function, an antioxidant effect, reduction of A beta production and neurotoxicity, reduced excitotoxicity, increased expression of the antiapoptotic factor bcl-2, and activation of mitogen activated protein kinase pathways. It is hypothesized that several of these neuroprotective mechanisms can be linked back to estrogen's ability to act as a potent chemical (i.e., electron-donating) antioxidant. Progesterone, on the other hand, has a membrane stabilizing effect that also serves to reduce the damage caused by lipid peroxidation. In addition, it may also provide neuroprotection by suppressing neuronal hyperexcitability. The following review will discuss experimental and clinical evidence for sex differences in outcome after acute brain trauma and stroke, review the evidence implicating estrogens and progestins as mediators of this neuroprotection following acute neurological injury, and finally, address the specific mechanisms by which these hormones may protect the brain following acute neurological injury.

Cyclin D-dependent kinases act as mitogen-responsive, rate-limiting controllers of G1 phase progression in mammalian cells. Two novel members of the mouse INK4 gene family, p19 and p18, that specifically inhibit the kinase activities of CDK4 and CDK6, but do not affect those of cyclin E-CDK2, cyclin A-CDK2, or cyclin B-CDC2, were isolated. Like the previously described human INK4 polypeptides, p16INK4a/MTS1 and p15INK4b/MTS2, mouse p19 and p18 are primarily composed of tandemly repeated ankyrin motifs, each ca. 32 amino acids in length, p19 and p18 bind directly to CDK4 and CDK6, whether untethered or in complexes with D cyclins, and can inhibit the activity of cyclin D-bound cyclin-dependent kinases (CDKs). Although neither protein interacts with D cyclins or displaces them from preassembled cyclin D-CDK complexes in vitro, both form complexes with CDKs at the expense of cyclins in vivo, suggesting that they may also interfere with cyclin-CDK assembly. In proliferating macrophages, p19 mRNA and protein are periodically expressed with a nadir in G1 phase and maximal synthesis during S phase, consistent with the possibility that INK4 proteins limit the activities of CDKs once cells exit G1 phase. However, introduction of a vector encoding p19 into mouse NIH 3T3 cells leads to constitutive p19 synthesis, inhibits cyclin D1-CDK4 activity in vivo, and induces G1 phase arrest.

Lowering of tumor interstitial hypertension, which acts as a barrier for tumor transvascular transport, has been proposed as a general strategy to enhance tumor uptake and therapeutic effects of anticancer drugs. The tyrosine kinase platelet-derived growth factor (PDGF) beta-receptor is one mediator of tumor hypertension. The effects of PDGF antagonists on chemotherapy response were investigated in two tumor models that display PDGF receptor expression restricted to the tumor stroma, and in which PDGF antagonists relieve tumor hypertension. Inhibitory PDGF aptamers and the PDGF receptor tyrosine kinase inhibitor STI571 enhanced the antitumor effect of Taxol on s.c. KAT-4 tumors in SCID mice. Treatment with only PDGF antagonists had no effect on tumor growth. Taxol uptake in tumors was increased by treatment with PDGF antagonists. Cotreatment with PDGF antagonists and Taxol was not associated with antiangiogenic effects, and PDGF antagonists did not enhance the Taxol effect on in vitro growth of KAT-4 cells. STI571 also increased the antitumor effects of 5-fluorouracil on s.c. PROb tumors in syngeneic BDIX rats, without increasing the effect of 5-fluorouracil on cultured PROb cells. Expression of PDGF receptors in tumor stroma, as well as tumor hypertension, occurs in most common solid tumors. Therefore, our results have implications for treatment regimens for large patient groups and merit clinical testing. In conclusion, our study identifies inhibition of PDGF signaling in tumor stroma as a novel, possibly general strategy for enhancement of the therapeutic effects chemotherapy.

Six Escherichia coli and 12 Klebsiella pneumoniae isolates from a single hospital expressed a common beta-lactamase with a pI of approximately 9.0 and were resistant to cefoxitin and cefotetan (MIC ranges, 64 to>> 128 and 16 to>> 128 micrograms/ml, respectively). Seventeen of the 18 strains produced multiple beta-lactamases. Most significantly, three K. pneumoniae strains were also resistant to imipenem (MICs, 8 to 32 micrograms/ml). Spectrophotometric beta-lactamase assays with purified enzyme indicated hydrolysis of cephamycins, in addition to cephaloridine and benzylpenicillin. The 4ene encoding the pI 9.0 beta-lactamase (designated ACT-1 for AmpC type) was cloned and sequenced, which revealed an ampC-type beta-lactamase gene that originated from Enterobacter cloacae and that had 86% sequence homology to the P99 beta-lactamase and 94% homology to the partial sequence of MIR-1. Southern blotting revealed that the gene encoding ACT-1 was on a large plasmid in some of the K. pneumoniae strains as well as on the chromosomes of all of the strains, suggesting that the gene is located on an easily mobilized element. Outer membrane protein profiles of the K. pneumoniae strains revealed that the three imipenem-resistant strains were lacking a major outer membrane protein of approximately 42 kDa which was present in the imipenem-susceptible strains. ACT-1 is the first plasmid-mediated AmpC-type beta-lactamase derived from Enterobacter which has been completely sequenced. This work demonstrates that in addition to resistance to cephamycins, imipenem resistance can occur in K. pneumoniae when a high level of the ACT-1 beta-lactamase is produced in combination with the loss of a major outer membrane protein.

Members of the TGF-beta superfamily of signalling molecules work by activating transmembrane receptors with phosphorylating activity (serine-threonine kinase receptors); these in turn phosphorylate and activate SMADs, a class of signal transducers. Activins are growth factors that act primarily through Smad2, possibly in partnership with Smad4, which forms heteromeric complexes with different ligand-specific SMADs after activation. In frog embryos, Smad2 participates in an activin-responsive factor (ARF), which then binds to a promoter element of the Mix.2 gene. The principal DNA-binding component of ARF is FAST-1, a transcription factor with a novel winged-helix structure. We now report that Smad4 is present in ARF, and that FAST-1, Smad4 and Smad2 co-immunoprecipitate in a ligand-regulated fashion. We have mapped the site of interaction between FAST-1 and Smad2/Smad4 to a novel carboxy-terminal domain of FAST-1, and find that overexpression of this domain specifically inhibits activin signalling. In a yeast two-hybrid assay, the FAST-1 carboxy terminus interacts with Smad2 but not Smad4. Deletion mutants of the FAST-1 carboxy terminus that still participate in ligand-regulated Smad2 binding no longer associated with Smad4 or ARF. These results indicate that Smad4 stabilizes a ligand-stimulated Smad2-FAST-1 complex as an active DNA-binding factor.

Interleukin-6 (IL-6, also known as B-cell stimulatory factor 2/interferon beta 2) was previously shown to support the proliferation of granulocyte/macrophage progenitors and indirectly support the formation of multilineage and blast cell colonies in cultures of spleen cells from normal mice. We report here that IL-3 and IL-6 act synergistically in support of the proliferation of murine multipotential progenitors in culture. The time course of total colony formation by spleen cells isolated from mice 4 days after injection of 5-fluorouracil (150 mg/kg) was significantly shortened in cultures containing both lymphokines relative to cultures supported by either of the two factors. Serial observations (mapping) of individual blast cell colonies in culture revealed that blast cell colonies emerged after random time intervals in the presence of IL-3. The average time of appearance in IL-6 alone was somewhat delayed, and in cultures containing both factors the appearance of multilineage blast cell colonies was significantly hastened relative to cultures grown in the presence of the individual lymphokines. In cultures of day-2 post-5-fluorouracil bone marrow cells, IL-6 failed to support colony formation; IL-3 alone supported the formation of a few granulocyte/macrophage colonies, but the combination of factors acted synergistically to yield multilineage and a variety of other types of colonies. In this system, IL-1 alpha also acted synergistically with IL-3, but the effect was smaller, and no multilineage colonies were seen. Together these results indicate that IL-3 and IL-6 act synergistically to support the proliferation of hemopoietic progenitors and that at least part of the effect results from a decrease in the G0 period of the individual stem cells.

The reproductive development of alligators from a contaminated and a control lake in central Florida was examined. Lake Apopka is adjacent to an EPA Superfund site, listed due to an extensive spill of dicofol and DDT or its metabolites. These compounds can act as estrogens. Contaminants in the lake also have been derived from extensive agricultural activities around the lake that continue today and a sewage treatment facility associated with the city of Winter Garden, Florida. We examined the hypothesis that an estrogenic contaminant has caused the current failure in recruitment of alligators on Lake Apopka. Supporting data include the following: At 6 months of age, female alligators from Lake Apopka had plasma estradiol-17 beta concentrations almost two times greater than normal females from the control lake, Lake Woodruff. The Apopka females exhibited abnormal ovarian morphology with large numbers of polyovular follicles and polynuclear oocytes. Male juvenile alligators had significantly depressed plasma testosterone concentrations comparable to levels observed in normal Lake Woodruff females but more than three times lower than normal Lake Woodruff males. Additionally, males from Lake Apopka had poorly organized testes and abnormally small phalli. The differences between lakes and sexes in plasma hormone concentrations of juvenile alligators remain even after stimulation with luteinizing hormone. Our data suggest that the gonads of juveniles from Lake Apopka have been permanently modified in ovo, so that normal steroidogenesis is not possible, and thus normal sexual maturation is unlikely.

OBJECTIVE

To understand the current status of knowledge in the basic field of polarized specific immune responses mediated by CD4+ T helper (Th) lymphocytes, based on their profile of cytokine production (type 1 or Th1 and type 2 or Th2).

METHODS

Relevant articles and publications from the medical literature, especially review articles dealing with properties, mechanisms of polarization, transcription regulatory factors, and role in different human pathophysiological conditions of Th1 and Th2 cells.

CONCLUSIONS

Th1 cells, which produce interferon (IFN)-gamma, interleukin (IL)-2 and tumor necrosis factor (TNF)-beta, evoke cell-mediated immunity and phagocyte-dependent inflammation. Th2 cells, which produce IL-4, IL-5, IL-6, IL-9, IL-10, and IL-13, evoke strong antibody responses (including those of the IgE class) and eosinophil accumulation, but inhibit several functions of phagocytic cells (phagocyte-independent inflammation). Both environmental and genetic factors act in concert to determine the Th1 or Th2 polarization. Further, Th1-dominated responses are involved in the pathogenesis of organ-specific autoimmune disorders, Crohn's disease, sarcoidosis, acute kidney allograft rejection, and some unexplained recurrent abortions. In contrast, allergen-specific Th2 responses are responsible for atopic disorders in genetically susceptible individuals. Further, Th2-dominated responses play a pathogenic role in both progressive systemic sclerosis and cryptogenic fibrosing alveolitis, and favor a more rapid evolution of HIV infection towards the full-blown disease. Finally, the Th1/Th2 paradigm can provide the basis for the development of new types of vaccines against infectious agents and of novel strategies for the therapy of allergic and autoimmune disorders.

The status of the 66-kDa human estrogen receptor-alpha (hER-alpha66) is a critical determinant in the assessment of the prognosis and in the design of treatment strategies of human breast cancer. Recently, we cloned the cDNA of an alternatively spliced variant of hER-alpha66, termed hER-alpha36; the predicted protein lacks both transcriptional activation domains of hER-alpha66 but retains its DNA-binding domain, partial dimerization, and ligand-binding domains and three potential myristoylation sites located near the N terminus. These findings thus predict that hER-alpha36 functions very differently from hER-alpha66 in response to estrogen signaling. We now demonstrate that hER-alpha36 inhibits the estrogen-dependent and estrogen-independent transactivation activities of hER-alpha66 and hER-beta. We further demonstrate that hER-alpha36 is predominantly associated with the plasma membrane where it transduces both estrogen- and antiestrogen-dependent activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase signaling pathway and stimulates cell growth. We conclude that hER-alpha36 is a predominantly membrane-based, unique alternatively spliced variant of hER-alpha66 that acts as a dominant-negative effector of both estrogen-dependent and estrogen-independent transactivation functions signaled through hER-alpha66 and ER-beta; it also transduces membrane-initiated estrogen-dependent activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase mitogenic signaling pathway. The estrogen and antiestrogen signaling pathways mediated by hER-alpha36 provide an alternative explanation for why some human breast cancers are resistant to and others are worsened by antiestrogen therapy; the data suggest that hER-alpha36 also may be an important marker to direct therapy in human breast cancers, and perhaps hER-alpha36 also may transduce estrogen-dependent signaling in other estrogen target tissues.

The SLC16 gene family has fourteen members. Four (SLC16A1, SLC16A3, SLC16A7, and SLC16A8) encode monocarboxylate transporters (MCT1, MCT4, MCT2, and MCT3, respectively) catalysing the proton-linked transport of monocarboxylates such as l-lactate, pyruvate and ketone bodies across the plasma membrane. SLC16A2 encodes a high affinity thyroid hormone transporter (MCT8) and SLC16A10 an aromatic amino acid transporter (TAT1). The substrates and roles of the remaining eight members are unknown. All family members are predicted to have 12 transmembrane helices (TMs) with intracellular C- and N-termini and a large intracellular loop between TMs 6 and 7. This topology has been confirmed for MCT1 and a three-dimensional structure has been modelled that suggests a plausible molecular mechanism. For correct plasma membrane expression and activity MCTs1-4, but not MCT8, require association with basigin or embigin; these are glycoproteins with a single TM and 2-3 extracellular immunoglobulin domains. SLC16 family members are involved in a wide range of metabolic pathways including energy metabolism of the brain, skeletal muscle, heart and tumour cells, gluconeogenesis, T-lymphocyte activation, bowel metabolism, spermatogenesis, pancreatic β-cell malfunction, thyroid hormone metabolism, and drug transport. MCTs 1-4 have distinct properties, tissue distribution and subcellular localisation that are appropriate for these metabolic roles. Their potential as pharmacological targets has been recognised with the discovery of potent and specific MCT1 inhibitors that act as immunosuppressant drugs by preventing proliferation of T-lymphocytes. It is suggested that the development of other drugs specifically targeting different MCT isoforms may provide a novel approach to cancer chemotherapy.

Tcf/Lef family transcription factors are the downstream effectors of the Wingless/Wnt signal transduction pathway. Upon Wingless/Wnt signalling, beta-catenin translocates to the nucleus, interacts with Tcf (1-3) and thus activates transcription of target genes (4,5). Tcf factors also interact with members of the Groucho (Grg/TLE) family of transcriptional co-repressors (6). We have now tested all known mammalian Groucho family members for their ability to interact specifically with individual Tcf/Lef family members. Transcriptional activation by any Tcf could be repressed by Grg-1, Grg-2/TLE-2, Grg-3 and Grg-4 in a reporter assay. Specific interactions between Tcf and Grg proteins may be achieved in vivo by tissue- or cell type-limited expression. To address this, we determined the expression of all Tcf and Grg/TLE family members in a panel of cell lines. Within any cell line, several Tcfs and TLEs are co-expressed. Thus, redundancy in Tcf/Grg interactions appears to be the rule. The 'long' Groucho family members containing five domains are repressors of Tcf-mediated transactivation, whereas Grg-5, which only contains the first two domains, acts as a de-repressor. As previously shown for Drosophila Groucho, we show that long Grg proteins interact with histone deacetylase-1. Although Grg-5 contains the GP homology domain that mediates HDAC binding in long Grg proteins, Grg-5 fails to bind this co-repressor, explaining how it can de-repress transcription.