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Biomedical subjects

S Frank

Publications and source records attributed to S Frank.

At least 37 records · Page 2Linked to original sources
PubMed2001

Urinary excretion of apolipoprotein(a): relation to other plasma proteins.

The atherogenic lipoprotein Lp(a) consists of an LDL-like core and apo(a), linked to apoB via a thiol bridge. Apo(a) fragments ranging in size from 60 to 220 kDa are excreted into urine and the excretion rate correlates significantly with the plasma levels of Lp(a). In order to study the interrelationship of apo(a) secretion with that of other plasma proteins, urinary apo(a) and protein secretion of five probands were followed for 24 h at different urinary densities. The excretion rate of apo(a) fragments, despite their high molecular weight, was highest, followed by apoD, orosomucoid, albumin and beta(2)-glycoprotein-I (beta2-GI) and plasminogen (1.58, 0.87, 0.095, 0.027, 0.013 and <0.001%/day, respectively). There was a highly significant correlation between apo(a), apoD and beta2-GI concentrations but not with albumin and orosomucoid concentrations in urine. The only protein that was fragmented in urine was apo(a) while the other proteins had molecular weights comparable to those in plasma. We conclude that a previously suggested fragmentation of apo(a) by the kidney is not a rate-limiting step in its excretion. Since plasminogen, another kringle-IV-containing plasma compound, and fragments thereof, are undetectable in urine under identical experimental conditions, it is very unlikely that the characteristic kringle structure is responsible for the high excretion rate of apo(a).

Adult↗
PubMed2001

The role of dynamin-related protein 1, a mediator of mitochondrial fission, in apoptosis.

In healthy cells, fusion and fission events participate in regulating mitochondrial morphology. Disintegration of the mitochondrial reticulum into multiple punctiform organelles during apoptosis led us to examine the role of Drp1, a dynamin-related protein that mediates outer mitochondrial membrane fission. Upon induction of apoptosis, Drp1 translocates from the cytosol to mitochondria, where it preferentially localizes to potential sites of organelle division. Inhibition of Drp1 by overexpression of a dominant-negative mutant counteracts the conversion to a punctiform mitochondrial phenotype, prevents the loss of the mitochondrial membrane potential and the release of cytochrome c, and reveals a reproducible swelling of the organelles. Remarkably, inhibition of Drp1 blocks cell death, implicating mitochondrial fission as an important step in apoptosis.

Animals↗
PubMed2001

Measuring change in quality of life in response to Helicobacter pylori eradication in peptic ulcer disease: the QOLRAD.

This study was designed to determine if a new condition-specific quality of life measure, the Quality Of Life in Reflux And Dyspepsia (QOLRAD), was responsive to clinical change over a four-week period when evaluating peptic ulcer disease patients undergoing Helicobacter pylori (Hp) eradication. A secondary aim was to evaluate QOLRAD's psychometric performance. Hp-positive patients with peptic ulcer disease were recruited from primary care and gastroenterology clinics (N = 155). QOLRAD scores improved significantly (P < 0.0001) and mean total score increase was 1.07 points on a 7-point scale among 111 subjects who responded to Hp eradication. Effect sizes were moderate for responders (0.64) and low for nonresponders (0.36). Score changes below 0.60 were associated with the smallest improvement in patient and physician condition rating; scores above 1.2 were associated with the greatest improvement. The QOLRAD demonstrates reliability and validity. Changes in QOLRAD score can be meaningfully related to clinical changes, aiding evaluation of health-related quality of life in peptic ulcer disease.

Dyspepsia↗
PubMed2001

The laminin-binding domain of agrin is structurally related to N-TIMP-1.

Agrin is the key organizer of postsynaptic differentiation at the neuromuscular junction. This organization activity requires the binding of agrin to the synaptic basal lamina. Binding is conferred by the N-terminal agrin (NtA) domain, which mediates a high-affinity interaction with the coiled coil domain of laminins. Here, we report the crystal structure of chicken NtA at 1.6 A resolution. The structure reveals that NtA harbors an oligosaccharide/oligonucleotide-binding fold with several possible sites for the interaction with different ligands. A high structural similarity of NtA with the protease inhibition domain in tissue inhibitor of metalloproteinases-1 (TIMP-1) supports the idea of additional functions of agrin besides synaptogenic activity.

Agrin↗
PubMed2001

Expressional regulation of angiopoietin-1 and -2 and the tie-1 and -2 receptor tyrosine kinases during cutaneous wound healing: a comparative study of normal and impaired repair.

It has become evident that a closely regulated presence of vascular endothelial growth factor (VEGF) and angiopoietin (Ang) factors determines the fate of blood vessel formation during angiogenesis. As angiogenesis is central to a normal wound-healing process, we investigated the regulation of Ang-1 and -2 and the related tyrosine kinase with immunoglobulin and epidermal growth factor homology (Tie)-1 and -2 receptors during normal repair in Balb/c mice and diabetes-impaired wound healing conditions in genetically diabetic (db/db) mice. For both normal and impaired healing conditions, we observed a constitutive expression of Ang-1, which was paralleled by an increase of Ang-2 upon injury. Whereas the observed Ang-2 expression declines from Day 7 after injury in control mice, diabetic-impaired healing was characterized by still increasing amounts of Ang-2 at these time points. Furthermore, Tie-1 was strongly induced during repair with a prolonged expression in diabetic mice, whereas Tie-2 expression was constitutive during normal repair but completely absent in diabetes-impaired healing. The overexpression of Ang-2 in the presence of markedly reduced VEGF in wounds of diabetic mice was associated with a dramatic decrease in endothelial cell numbers compared with normal healing as assessed by analysis of the endothelium-specific markers CD31 and von Willebrand factor, whereas the lymphatic endothelium remained stable as determined by expression of VEGF receptor-3 (VEGFR-3/Flt-4).

Angiopoietin-1↗
PubMed2001

Adenovirus-mediated apo(a)-antisense-RNA expression efficiently inhibits apo(a) synthesis in vitro and in vivo.

Apo(a) is a very atherogenic plasma protein without apparent function, which is highly expressed in humans. The variation in plasma Lp(a) concentration among individuals is considerable. Approximately 10-15% of the white population exhibit plasma Lp(a) concentrations above the atherogenic cut-off value of approximately 30 mg/dl. Since there is currently no safe way of treating those patients with drugs, we have tested the possibility of interfering with apo(a) biosynthesis by adenovirus-mediated expression of antisense apo(a) mRNA comprising the 5' UTR, the signal sequence and the first three kringles of native apo(a). Transduction of rat hepatoma McA RH 7777 cells which stably expressed apo(a) with 18 kringle IV (KIV) domains with apo(a)-antisense adenovirus (AS-Ad) at multiplicity of infection (MOI) of 30 reduced apo(a) synthesis to 23% as compared with control cells. As apo(a) is not synthesized in laboratory animals, we induced biosynthesis of the N-terminal fragments of apo(a) in mice by adenovirus-mediated gene transfer. Cotransduction of these mice with AS-Ad, which expressed up to eight times higher amounts of apo(a) than stable transgenic apo(a) mice, led to an almost complete disappearance of apo(a) from plasma. We conclude that the proposed AS-construct is very efficient in interfering with apo(a) biosynthesis in vivo. The strategy of inducing the synthesis of a nonexpressed protein followed by knocking it out by AS technology may also be applicable to other systems.

Adenoviridae↗
PubMed2001

A novel keratinocyte mitogen: regulation of leptin and its functional receptor in skin repair.

Wound re-epithelialization represents a tissue movement that crucially participates in wound closure. Recently, we demonstrated that supplemented leptin improved re-epithelialization processes in leptin-deficient ob/ob mice. In this study we investigated regulation of the leptin system during normal repair in healthy animals. We found leptin to be present at the wound site during healing, although leptin levels were clearly reduced upon injury compared with uninvolved control skin. The functional leptin receptor subtype obRb was observed to be constitutively expressed in nonwounded skin. During early healing, the leptin receptor obRb was downregulated, but re-increased again from day 5 postwounding. Immunohistochemistry revealed that highly proliferative keratinocytes of the wound margin epithelia strongly expressed the functional leptin receptor subtype obRb. In vitro studies demonstrated that murine and human primary epidermal keratinocytes responded to exogenously added leptin with a proliferative response. Moreover, specificity of leptin-mediated mitogenic effects on primary keratinocytes could be shown by completely blocking leptin actions by a soluble, nonfunctional chimeric leptin receptor. Finally, we report that leptin, besides the recently described activation of the janus tyrosine kinase signal transducers, also activated extracellular signal-regulated kinase-controlled signaling pathways in primary keratinocytes.

Animals↗
PubMed2001

Role of various tissues in apo(a) fragmentation and excretion of fragments by the kidney.

BACKGROUND: Lipoprotein(a) [Lp(a)] is an atherothrombotic plasma lipoprotein with unknown function. Little is known about the catabolism of this lipoprotein, in particular the steps related to apolipoprotein(a) [apo(a)] fragmentation and excretion by the kidney. MATERIAL AND METHODS: High plasma levels (up to 9 mg dL(-1)) of the N-terminal fragment of apo(a) were expressed in mice by adenovirus mediated gene transfer. Plasma of such N-apo(a) mice was injected into acceptor mice and the fragmentation and urinary secretion of N-apo(a) were followed by immunochemical techniques. RESULTS: Mice transduced with N-Ad expressed apo(a)-fragments with 3-11 kringle-IV (KIV) repeats. Injection of N-apo(a)-plasma from donor mice into acceptor mice resulted in fragmentation of N-apo(a)s with 3-11 KIVs yielding smaller peptides down to 2 KIVs. Secretion of N-apo(a)-fragments with 2 to maximally 6 KIVs into urine occurred as early as 2 min after injection. Immunohistochemical studies of kidney suggested filtration as a mechanism of apo(a)-fragment excretion. When N-apo(a) was incubated in vitro with various tissues from perfused mice, skeletal muscle and kidney followed by liver and spleen contributed to fragmentation. Tissues from unperfused organs, or the addition of normal mouse plasma, caused marked reduction in N-apo(a) fragmentation. EDTA, and not aprotinin or leupeptin, prevented apo(a) cleavage. CONCLUSION: Here we provide evidence that apo(a) is cleaved by metalloproteinases located on skeletal muscle, kidney and other organs. Small apo(a)-fragments up to a size of 6 KIVs are excreted into urine, yet a major portion of apo(a) fragments is removed from circulation extrarenally.

Adenoviridae↗
PubMed2001

Scavenger receptor class B, type I is expressed in porcine brain capillary endothelial cells and contributes to selective uptake of HDL-associated vitamin E.

It is clearly established that an efficient supply to the brain of alpha-tocopherol (alphaTocH), the most biologically active member of the vitamin E family, is of the utmost importance for proper neurological functioning. Although the mechanism of uptake of alphaTocH into cells constituting the blood-brain barrier (BBB) is obscure, we previously demonstrated that high-density lipoprotein (HDL) plays a major role in the supply of alphaTocH to porcine brain capillary endothelial cells (pBCECs). Here we studied whether a porcine analogue of human and rodent scavenger receptor class B, type I mediates selective (without concomitant lipoprotein particle internalization) uptake of HDL-associated alphaTocH in a similar manner to that described for HDL-associated cholesteryl esters (CEs). In agreement with this hypothesis we observed that a major proportion of alphaTocH uptake by pBCECs occurred by selective uptake, exceeding HDL3 holoparticle uptake by up to 13-fold. The observation that selective uptake of HDL-associated CE exceeded HDL3 holoparticle up to fourfold suggested that a porcine analogue of SR-BI (pSR-BI) may be involved in lipid uptake at the BBB. In line with the observation of selective lipid uptake, RT-PCR and northern and western blot analyses revealed the presence of pSR-BI in cells constituting the BBB. Adenovirus-mediated overexpression of the human analogue of SR-BI (hSR-BI) in pBCECs resulted in a fourfold increase in selective HDL-associated alphaTocH uptake. In accordance with the proposed function of SR-BI, selective HDL-CE uptake was increased sixfold in Chinese hamster ovary cells stably transfected with murine SR-BI (mSR-BI). Most importantly stable mSR-BI overexpression mediated a twofold increase in HDL-associated [14C]alphaTocH selective uptake in comparison with control cells. In line with tracer experiments, mass transfer studies with unlabelled lipoproteins revealed that mSR-BI overexpression resulted in a twofold increase in endogenous HDL3-associated alphaTocH uptake. The results of this study indicate that SR-BI promotes the uptake of HDL-associated alphaTocH into cells constituting the BBB and plays an important role during the supply of the CNS with this indispensable micronutrient.

Animals↗
PubMed2001

Isolation and characterization of the human forkhead gene FOXQ1.

We have isolated a human genomic and cDNA clone that encodes a protein of 403 amino acids and belongs to the family of the FOX transcription factors (previously called HNF-3/forkhead transcription factors). The 2.7-kb transcript of the human FOXQ1 gene is expressed predominantly in the stomach, trachea, bladder and salivary gland. Additionally, overexpression of human FOXQ1 was shown in colorectal adenocarcinoma and lung carcinoma cell lines. The FOXQ1 gene is located on chromosome 6p23-25. Databank analysis shows 82% homology with the mouse Foxq1 gene (formerly Hfh-1L) and with a revised sequence of the rat FoxQ1 gene (formerly HFH-1). The DNA-binding motif, named HNF-3/forkhead domain, is well conserved, showing 100% identity in human, mouse, and rat. The human protein sequence contains two putative transcriptional activation domains, which share a high amino acid identity with the corresponding mouse and rat domains.

Amino Acid Sequence↗
PubMed2001

Papillomavirus type 16 oncogenes downregulate expression of interferon-responsive genes and upregulate proliferation-associated and NF-kappaB-responsive genes in cervical keratinocytes.

Infection with high-risk human papillomaviruses (HPV) is a major risk factor for development of cervical cancer. Expression of the HPV E6 and E7 oncoproteins increases in differentiating keratinocytes, resulting in inactivation of the p53 and retinoblastoma proteins, two important transcriptional regulators. We used cDNA microarrays to examine global alterations in gene expression in differentiating cervical keratinocytes after infection with retroviruses encoding HPV type 16 (HPV-16) E6 and E7. Expression of 80 cellular genes (approximately 4% of the genes on the array) was altered reproducibly by E6 and/or E7. Cluster analysis classified these genes into three functional groups: (i) interferon (IFN)-responsive genes, (ii) genes stimulated by NF-kappaB, and (iii) genes regulated in cell cycle progression and DNA synthesis. HPV-16 E6 or a dominant negative p53 protein downregulated multiple IFN-responsive genes. E6 decreased expression of IFN-alpha and -beta, downregulated nuclear STAT-1 protein, and decreased binding of STAT-1 to the IFN-stimulated response element. E7 alone was less effective; however, coexpression of E6 and E7 downregulated IFN-responsive genes more efficiently than E6. The HPV-16 E6 protein also stimulated expression of multiple genes known to be inducible by NF-kappaB and AP-1. E6 enhanced expression of functional components of the NF-kappaB signal pathway, including p50, NIK, and TRAF-interacting protein, and increased binding to NF-kappaB and AP-1 DNA consensus binding sites. Secretion of interleukin-8, RANTES, macrophage inflammatory protein 1alpha, and 10-kappaDa IFN-gamma-inducible protein were increased in differentiating keratinocytes by E6. Thus, high-level expression of the HPV-16 E6 protein in differentiating keratinocytes directly alters expression of genes that influence host resistance to infection and immune function.

Cell Cycle↗
PubMed2001

Molecular cloning of ILP-2, a novel member of the inhibitor of apoptosis protein family.

Inhibitor of apoptosis protein (IAP)-like protein-1 (ILP-1) (also known as X-linked IAP [XIAP] and mammalian IAP homolog A [MIHA]) is a potent inhibitor of apoptosis and exerts its effects, at least in part, by the direct association with and inhibition of specific caspases. Here, we describe the molecular cloning and characterization of a human gene related to ILP-1, termed ILP-2. Despite high homology to ILP-1, ILP-2 is encoded by a distinct gene, which in normal tissues is expressed solely in testis. In contrast to ILP-1, overexpression of ILP-2 had no protective effect on apoptosis mediated by Fas (also known as CD95) or tumor necrosis factor. However, ILP-2 potently inhibited apoptosis induced by overexpression of Bax or by coexpression of caspase 9 with Apaf-1, and preincubation of cytosolic extracts with ILP-2 abrogated caspase activation in vitro. A processed form of caspase 9 could be coprecipitated with ILP-2 from cells, suggesting a physical interaction between ILP-2 and caspase 9. Thus, ILP-2 is a novel IAP family member with restricted specificity for caspase 9.

Amino Acid Sequence↗
PubMed2001

Impact of apolipoprotein(a) on in vitro angiogenesis.

Angiostatin, which consists of the kringle I-IV domains of plasminogen and which is secreted into urine, is an efficient inhibitor of angiogenesis and tumor growth. Because N-terminal apolipoprotein(a) [apo(a)] fragments, which also contain several types of kringle IV domains, are found in urine as well, we evaluated the potential angiostatic properties of these urinary apo(a) fragments and of a recombinant form of apo(a) [r-apo(a)]. We used human microvascular endothelial cell (hMVEC)-based in vitro assays of tube formation in 3-dimensional fibrin matrixes. Purified urinary apo(a) fragments or r-apo(a) inhibited the basic fibroblast growth factor/tumor necrosis factor-alpha-induced formation of capillary-like structures. At concentrations varying from 0.2 to 10 microgram/mL, urinary apo(a) fragments inhibited tube formation by as much as 70%, whereas there was complete inhibition by r-apo(a). The highest concentrations of both inhibitors also reduced urokinase plasminogen activator production of basic fibroblast growth factor-induced hMVEC proliferation. The inhibitors had no effect on plasminogen activator inhibitor-1 expression. If our in vitro model for angiogenesis is valid for the in vivo situation as well, our data point toward the possibility that apo(a) may also be physiologically operative in modulating angiogenesis, as the concentration of free apo(a) found in humans exceeds that tested herein.

Angiogenesis Inhibitors↗
PubMed2001

License to kill tumors: how much hope is justified for trail?

In 1995, a new cytokine termed TRAIL (tumor necrosis factor--related apoptosis-inducing ligand) was discovered and demonstrated to selectively induce programmed cell death in transformed cell lines. Preclinical cytotoxicity studies in mice and nonhuman primates have produced promising results by demonstrating that TRAIL exerts potent tumoricidal activity but lacks severe toxicity towards normal tissues making it a potentially ideal candidate for cancer therapy. This article reviews aspects of our current understanding of TRAIL signaling pathways and summarizes how this knowledge is currently being translated into TRAIL-based tumor-selective therapeutic strategies.

Antineoplastic Agents↗
PubMed2001

Macrophage-derived heme-oxygenase-1: expression, regulation, and possible functions in skin repair.

BACKGROUND: Expression and enzymatic activity of heme oxygenase (HO) has been implicated in the development, as well as in the resolution, of inflammatory conditions. Because inflammation is central to tissue repair, we investigated the presence and potential functions of HO in an excisional model of normal and diabetes-impaired wound repair in mice. MATERIALS AND METHODS: Expression of HO-1 during cutaneous healing was analyzed by RNase protection assay, Western blot, and immunohistochemical techniques in a murine model of excisional repair. Furthermore, we determined HO-1-dependent release of proinflammatory cytokines from RAW 264.7 macrophages by enzyme-linked immunosorbent assay (ELISA). RESULTS: Upon injury, we observed a rapid and strong increase in HO-1 mRNA and protein levels at the wound site. By contrast to normal repair, late stages of diabetes-impaired repair were associated with elevated HO-1 expression. Besides a few keratinocytes of the hyperproliferative epithelium, immunohistochemistry revealed infiltrating macrophages as the predominant and major source of HO-1 at the wound site. In vitro studies demonstrated the potency of exogenous and also endogenous nitric oxide (NO) to strongly induce HO-1 expression in RAW 264.7 macrophages. However, L-NIL-mediated enzymatic inhibition of inducible NO-synthase (iNOS) at the wound site in vivo was not paralleled by decreased HO-1 levels. In vitro inhibition of HO-1 enzymatic activity by tin protoporphyrin IX (SnPPIX) in RAW 264.7 macrophages markedly attenuated tumor necrosis factor-alpha (TNF-alpha), but strongly increased interleukin-1beta (IL-1beta) release in RAW 264.7 macrophages in vitro. CONCLUSIONS: The observed injury-mediated increase in HO-1 mRNA and protein at the wound site was due to infiltrating HO-1 expressing monocytic cells. Macrophage-derived HO-1 expression was not under regulatory control by NO in skin repair. We provide evidence that HO-1 might exert a regulatory role in macrophage-derived cytokine release.

Animals↗
PubMed2000

An intrahelical salt bridge within the trigger site stabilizes the GCN4 leucine zipper.

We previously reported that a helical trigger segment within the GCN4 leucine zipper monomer is indispensable for the formation of its parallel two-stranded coiled coil. Here, we demonstrate that the intrinsic secondary structure of the trigger site is largely stabilized by an intrahelical salt bridge. Removal of this surface salt bridge by a single amino acid mutation induced only minor changes in the backbone structure of the GCN4 leucine zipper dimer as verified by nuclear magnetic resonance. The mutation, however, substantially destabilized the dimeric structure. These findings support the proposed hierarchic folding mechanism of the GCN4 coiled coil in which local helix formation within the trigger segment precedes dimerization.

Alanine↗
PubMed2000

Keratinocyte-derived chemotactic cytokines: expressional modulation by nitric oxide in vitro and during cutaneous wound repair in vivo.

Inhibition of inducible nitric oxide-synthase (iNOS) enzymatic activity during cutaneous wound repair leads to severely impaired tissue regeneration. To assess whether disturbed leukocyte infiltration might participate in impaired repair, we determined expressional kinetics of neutrophil-attracting macrophage inflammatory protein-2 (MIP-2), and monocyte-attracting macrophage chemoattractant protein-1 (MCP-1) using an excisional wound healing model in mice. MCP-1 was induced in epithelial keratinocytes upon wounding, and our data indicate that NO serves a negative regulatory role for MCP-1 expression in vivo, as clearly reduced numbers of wound margin keratinocytes associated with NO-deficient repair compensate for high MCP-1 expression levels observed during normal healing. MIP-2 expression was restricted to hair follicles which were not reduced in number during NO-deficient repair. In vitro studies confirmed a regulatory role of NO for keratinocyte-derived chemokine expression, as NO attenuated IL-1beta- and TNF-alpha-induced MCP-1 mRNA expression, whereas NO augmented IL-1beta-induced IL-8 (functional human homolog to murine MIP-2) mRNA expression in the human keratinocyte cell line HaCaT.

Animals↗