gebhart/patrepeval huggingface dataset:
Development of a novel mammalian cell surface antibody display platform (2010)

Antibody display systems have been successfully applied to screen, select and characterize antibody fragments. These systems typically use prokaryotic organisms such as phage and bacteria or lower eukaryotic organisms, such as yeast. These organisms possess either no or different post-translational modification functions from mammalian cells and prefer to display small antibody fragments instead of full-length IgGs. We report here a novel mammalian cell-based antibody display platform that displays full-length functional antibodies on the surface of mammalian cells. Through recombinase-mediated DNA integration, each host cell contains one copy of the gene of interest in the genome. Utilizing a hot-spot integration site, the expression levels of the gene of interest are high and comparable between clones, ensuring a high signal to noise ratio. Coupled with fluorescence-activated cell sorting (FACS) technology, our platform is high throughput and can distinguish antibodies with very high antigen binding affinities directly on the cell surface. Single-round FACS can enrich high affinity antibodies by more than 500 fold. Antibodies with significantly improved neutralizing activity have been identified from a randomly mutagenized library, demonstrating the power of this platform in screening and selecting antibody therapeutics.
Large Amounts of Cell-free Fetal DNA Are Present in Amniotic Fluid (2001)

Until recently, the focus of our research in the noninvasive diagnosis of Down syndrome was the genetic analysis of intact fetal nucleated erythrocytes from the circulation of pregnant women (1). The initial step in the physical isolation of these target cells involves the layering of diluted maternal blood on a density gradient (2). Before 1997, we routinely discarded the plasma layer of the density gradient, unaware of the fact that it could contain nucleic acids. In 1997, prompted by reports of large quantities of tumor-specific DNA sequences in the plasma and serum of cancer patients (3)(4)(5), Lo et al. (6) demonstrated the presence of male fetal DNA sequences in the serum and plasma of pregnant women. Subsequently, this same group extended their observations by quantifying the fetal DNA in maternal plasma (7) and studying its kinetics and physiology (8).

The detection and/or quantification of fetal DNA sequences in maternal plasma have been used for a variety of clinical applications, including diagnosis of gender, Rhesus D genotype, single gene disorders, aneuploidy, and preeclampsia [reviewed in Refs. (9)(10)]. More recently, it has also been suggested that the genetic material in the plasma consists of a continuum of intact and apoptotic cells as well as cell-free DNA (11)(12). Fetal DNA, however, appears to exist predominantly in a cell-free form in the maternal plasma (13). Despite advances in the clinical applications of this technology, to date not much is known about the tissue of origin of the cell-free fetal DNA and how it is metabolized in the pregnant woman.

Botezatu et al. (14) demonstrated that male DNA sequences could be found in the urine of women who received a blood transfusion from a male donor, as well as women who were pregnant with a male …
Tocopherol‐mediated peroxidation of lipoproteins: implications for vitamin E as a potential antiatherogenic supplement (1999)

The 'oxidation theory' of atherosclerosis proposes that oxidation of low density lipoprotein (LDL) contributes to atherogenesis. Although little direct evidence for a causative role of 'oxidized LDL' in atherogenesis exists, several studies show that, in vitro, oxidized LDL exhibits potentially proatherogenic activities and lipoproteins isolated from atherosclerotic lesions are oxidized. As a consequence, the molecular mechanisms of LDL oxidation and the actions of alpha-tocopherol (alpha-TOH, vitamin E), the major lipid-soluble lipoprotein antioxidant, have been studied in detail. Based on the known antioxidant action of alpha-TOH and epidemiological evidence, vitamin E is generally considered to be beneficial in coronary artery disease. However, intervention studies overall show a null effect of vitamin E on atherosclerosis. This confounding outcome can be rationalized by the recently discovered diverse role for alpha-TOH in lipoprotein oxidation; that is, alpha-TOH displays neutral, anti-, or, indeed, pro-oxidant activity under various conditions. This review describes the latter, novel action of alpha-TOH, termed tocopherol-mediated peroxidation, and discusses the benefits of vitamin E supplementation alone or together with other antioxidants that work in concert with alpha-TOH in ameliorating lipoprotein lipid peroxidation in the artery wall and, hence, atherosclerosis.
Limitations of Conventional Methods of Self-Monitoring of Blood Glucose (2001)

Despite excellent HbA(1c) levels and target preprandial glucose levels, children often experience nocturnal hypoglycemia and postprandial hyperglycemia that are not evident with routine monitoring. Repeated use of the CGMS may provide a means to optimize basal and bolus insulin replacement in patients with type 1 diabetes.
Engineering the Growth Factor Microenvironment with Fibronectin Domains to Promote Wound and Bone Tissue Healing (2011)

Although growth factors naturally exert their morphogenetic influences within the context of the extracellular matrix microenvironment, the interactions among growth factors, their receptors, and other extracellular matrix components are typically ignored in clinical delivery of growth factors. We present an approach for engineering the cellular microenvironment to greatly accentuate the effects of vascular endothelial growth factor-A (VEGF-A) and platelet-derived growth factor-BB (PDGF-BB) for skin repair, and of bone morphogenetic protein-2 (BMP-2) and PDGF-BB for bone repair. A multifunctional recombinant fragment of fibronectin (FN) was engineered to comprise (i) a factor XIIIa substrate fibrin-binding sequence, (ii) the 9th to 10th type III FN repeat (FN III9-10) containing the major integrin-binding domain, and (iii) the 12th to 14th type III FN repeat (FN III12-14), which binds growth factors promiscuously, including VEGF-A165, PDGF-BB, and BMP-2. We show potent synergistic signaling and morphogenesis between α5β1 integrin and the growth factor receptors, but only when FN III9-10 and FN III12-14 are proximally presented in the same polypeptide chain (FN III9-10/12-14). The multifunctional FN III9-10/12-14 greatly enhanced the regenerative effects of the growth factors in vivo in a diabetic mouse model of chronic wounds (primarily through an angiogenic mechanism) and in a rat model of critical-size bone defects (through a mesenchymal stem cell recruitment mechanism) at doses where the growth factors delivered within fibrin only had no significant effects.
Attomole level protein sequencing by Edman degradation coupled with accelerator mass spectrometry (2001)

Edman degradation remains the primary method for determining the sequence of proteins. In this study, accelerator mass spectrometry was used to determine the N-terminal sequence of glutathione S-transferase at the attomole level with zeptomole precision using a tracer of (14)C. The transgenic transferase was labeled by growing transformed Escherichia coli on [(14)C]glucose and purified by microaffinity chromatography. An internal standard of peptides on a solid phase synthesized to release approximately equal amounts of all known amino acids with each cycle were found to increase yield of gas phase sequencing reactions and subsequent semimicrobore HPLC as did a lactoglobulin carrier. This method is applicable to the sequencing of proteins from cell culture and illustrates a path to more general methods for determining N-terminal sequences with high sensitivity.
Aggresomes: A Cellular Response to Misfolded Proteins (1998)

Intracellular deposition of misfolded protein aggregates into ubiquitin-rich cytoplasmic inclusions is linked to the pathogenesis of many diseases. Why these aggregates form despite the existence of cellular machinery to recognize and degrade misfolded protein and how they are delivered to cytoplasmic inclusions are not known. We have investigated the intracellular fate of cystic fibrosis transmembrane conductance regulator (CFTR), an inefficiently folded integral membrane protein which is degraded by the cytoplasmic ubiquitin-proteasome pathway. Overexpression or inhibition of proteasome activity in transfected human embryonic kidney or Chinese hamster ovary cells led to the accumulation of stable, high molecular weight, detergent-insoluble, multiubiquitinated forms of CFTR. Using immunofluorescence and transmission electron microscopy with immunogold labeling, we demonstrate that undegraded CFTR molecules accumulate at a distinct pericentriolar structure which we have termed the aggresome. Aggresome formation is accompanied by redistribution of the intermediate filament protein vimentin to form a cage surrounding a pericentriolar core of aggregated, ubiquitinated protein. Disruption of microtubules blocks the formation of aggresomes. Similarly, inhibition of proteasome function also prevented the degradation of unassembled presenilin-1 molecules leading to their aggregation and deposition in aggresomes. These data lead us to propose that aggresome formation is a general response of cells which occurs when the capacity of the proteasome is exceeded by the production of aggregation-prone misfolded proteins.
Expression of MMP-7, MMP-9, TIMP-1 and TIMP-2 mRNA in lung tissue of patients with non-small cell lung cancer (NSCLC) and benign pulmonary disease. (2009)

Significantly higher expression of MMP-7 and MMP-9 in tumor tissue than in the surrounding tissue or in benign lung disease tissue supports the notion of an important role of these metalloproteinases in the growth of lung carcinoma. TIMP-1 expression is increased only in carcinoma, but not in benign lung disease.
trans-Activation of the human T-cell leukemia virus long terminal repeat correlates with expression of the x-lor protein (1985)

Cell lines established directly from adult T-cell leukemia-lymphoma patients or immortalized by human T-cell leukemia virus type I (HTLV-I) in vitro that do not produce complete HTLV virions were characterized both for the content of viral proteins and for the presence of trans-acting factors activating gene expression under the control of the HTLV long terminal repeat. The expression of the 42-kilodalton HTLV x-lor product correlated with trans-activation of the long terminal repeat. The implications of this study for understanding the role of the HTLV x-lor product in the initiation and maintenance of T-lymphocyte transformation are discussed.
Intramuscular injection of naked plasmid DNA encoding human preproinsulin gene in streptozotocin-diabetes mice results in a significant reduction of blood glucose level. (2003)

The insulin complement with gene therapy has been used as an experimental treatment for insulin dependent diabetes (IDDM). In the present study, we constructed naked plasmid DNA vector encoding recombinant human preproinsulin gene (pCMV-IN), and injected the plasmids (100 microg/mouse) intramuscularly combined with electroporation, to achieve the in vivo transfer of insulin gene in streptozotocin (STZ)-induced diabetic C57 mice. The expression of vector-derived insulin mRNA was detected with RT-PCR in transfected local skeletal muscles. The plasma insulin was elevated significantly in pCMV-IN injected diabetic C57 mice, which was complemented to the level similar to the intact normal control. The protein expression lasted for at least 35 days after the plasmid injection. Gene therapy with pCMV-IN plasmids considerably decreased the blood glucose level in STZ-induced diabetic mice from d 7 to d 35 by about 6 mmol/L. The gene therapy also reduced the mortality of severe diabetic mice significantly from 100% to 37% at the 6th week. Our results indicate that the direct intramuscular injection of naked plasmids encoding human preproinsulin gene achieves the effective expression of insulin. The restoration of insulin decreases blood glucose and increases the survival in severe diabetic mice. The gene therapy might be provided as a practical therapeutic approach to IDDM.
Tumour‐associated macrophages and melanoma tumourigenesis: integrating the complexity (2006)

When the body discovers a tumour cell (foreign antigen), several kinds of mechanisms and cells operate in what is called an immune response. The latter has evolved into two mechanisms: non-specific immunity and specific immunity, which are closely linked to and influence each other. The former represents the first line of defence against neoplastic cells. The adaptive (specific) immunity is orchestrated by antigen-specific T and B lymphocytes. The effector cells of innate immunity include granulocytes, macrophages and natural killer cells. Among these cells, macrophages represent the most important part of innate immunity against tumours. Tumour-associated macrophages (TAMs) are important antigen-presenting cells and as such an understanding of their interactions with tumour cells gives insights into novel therapeutic strategies. In tumours, the effect of TAMs is the outcome of their two concomitantly competing interactions: tumour growth reduction and tumour growth promotion. The macrophage (TAMs) content of melanoma ranges from 0 to 30% and their density increases with increasing tumour thickness. The melanoma cells and TAMs seem to interact with each other through the release of soluble factors that either prevent or enhance tumour growth. For instance, syngeneic macrophages from tumour-bearing mice can inhibit melanoma growth in the nude mice more than the control macrophages. Alternatively, metastatic B16 melanoma cells can produce some macrophage cytotoxic substances that help tumour cells not only escape the host immunosurveillance system but also prevent distant metastasis. Together, these observations suggest opposing effects for these soluble factors in melanoma. To date, little is available in the literature about the interactions between TAMs and melanoma cells. This viewpoint not only tries to examine these interactions but also provides relevant speculations.
Niosomal methotrexate gel in the treatment of localized psoriasis: Phase I and phase II studies (2007)

These results suggest that niosomal methotrexate gel is more efficacious than placebo and marketed methotrexate gel.
An internalization signal in the simian immunodeficiency virus transmembrane protein cytoplasmic domain modulates expression of envelope glycoproteins on the cell surface. (1996)

A Tyr to Cys mutation at amino acid position 723 in the cytoplasmic domain of the simian immunodeficiency virus (SIV) transmembrane (TM) molecule has been shown to increase expression of envelope glycoproteins on the surface of infected cells. Here we show that Tyr-723 contributes to a sorting signal that directs the rapid endocytosis of viral glycoproteins from the plasma membrane via coated pits. On cells infected by SIVs with a Tyr at position 723, envelope glycoproteins were transiently expressed on the cell surface and then rapidly endocytosed. Similar findings were noted for envelope molecules expressed in the absence of other viral proteins. Immunoelectron microscopy demonstrated that these molecules were localized in patches on the cell surface and were frequently associated with coated pits. In contrast, envelope glycoproteins containing a Y723C mutation were diffusely distributed over the entire plasma membrane. To determine if an internalization signal was present in the SIV TM, chimeric molecules were constructed that contained the CD4 external and membrane spanning domains and a SIV TM cytoplasmic tail with a Tyr or other amino acids at SIV position 723. In Hela cells stably expressing these molecules, chimeras with a Tyr-723 were rapidly endocytosed, while chimeras containing other amino acids at position 723, including a Phe, were internalized at rates only slightly faster than a CD4 molecule that lacked a cytoplasmic domain. In addition, the biological effects of the internalization signal were evaluated in infectious viruses. A mutation that disrupted the signal and as a result, increased the level of viral envelope glycoprotein on infected cells, was associated with accelerated infection kinetics and increased cell fusion during viral replication. These results demonstrate that a Tyr-dependent motif in the SIV TM cytoplasmic domain can function as an internalization signal that can modulate expression of the viral envelope molecules on the cell surface and affect the biological properties of infectious viruses. The conservation of an analogous Tyr in all human and simian immunodeficiency viruses suggests that this signal may be present in other primate lentiviruses and could be important in the pathogenesis of these viruses in vivo.
Silicon micromachined microphone chip at Siemens (1999)

Applications ranging from hearing aids over communication to noise cancellation open up a high volume market for low-cost, batch producible and reliable microphones. To obey these conditions, a single-chip capacitive microphone has been developed at Siemens, utilizing a modified standard CMOS process with adjacent bulk micromachining. In a first step, the microphone is integrated with a source follower, enabling low output impedance of the signal. The technology allows for the future integration of advanced circuitry. The microphone consists of an acoustically sensitive polycrystalline silicon membrane and a highly perforated back-plate as the counter electrode. To achieve highly sensitive devices, special emphasis was given to the stress of the polycrystalline silicon membrane, which should be slightly tensile. Another key issue during the fabrication and in operation is to prevent stiction of the sensitive membrane. Since the overall chip size is below 3-mm side length, surface mounting in low-cost SMD packages is possible.
Characterization and growth factor requirements of SJL lymphomas. II. Interleukin 5 dependence of the in vitro cell line, cRCS‐X, and influence of other cytokines (1989)

Follicular B cell lymphomas of SJL mice [reticulum cell sarcoma (RCS)] are dependent on syngeneic Ly-1+,2- T cells for their growth. These T cells produce a number of lymphokines in response to stimulation with gamma-irradiated RCS cells including interleukin (IL) 2, interferon-gamma (IFN-gamma), IL4 and IL5, some of which may be required for growth of the tumor. Previous studies have shown that an RCS cell line, cRCS-X, can be maintained in vitro indefinitely, if the cultures are supplemented with gamma-irradiated lymph node (LN) cells or with a preparation of human B cell growth factor (BCGF). In the present studies, the growth requirements of this cell line were analyzed in more detail in short-term assays of both [3H]thymidine incorporation and colony formation in agarose. Recombinant murine IL5 cause dose-dependent proliferation of cRCS-X cells similar to that induced with BCGF. The level of colony formation by cRCS-X cells induced by optimal concentrations of BCGF was not increased further by the addition of IL5, suggesting that the two factors act via a common mechanism. IL1 and IFN-gamma each enhanced cRCS-X proliferation induced by BCGF or IL5 in both assays. The effects of IL1 plus BCGF, IFN-gamma plus BCGF, and IL1 plus IL5 were clearly synergistic. Preincubation of cRCS-X cells with IL1 enhanced their ability to proliferate in response to BCGF or IL5 in [3H]thymidine incorporation assays, but the reverse sequence of cytokine addition showed no effect of IL1. No such effect was seen with IFN-gamma. Indeed, IL1 and IFN-gamma appeared to affect BCGF-induced cRCS-X growth by different mechanisms and their combined effects were greater than that of IL1 or IFN-gamma added separately. None of the other cytokines studied, including IL2, IL3, IL4, IL6, tumor necrosis factor-alpha, granulocyte monocyte-colony-stimulating factor or transforming growth factor-beta, had any detectable effect on cRCS-X cells, either alone or in combination with BCGF or IL5. Like IL5, SJL gamma-irradiated LN cells induced cRCS-X colony-forming units (CFU) in a dose-dependent manner. IL1, or the combination of IL1 plus IFN-gamma, clearly synergized with LN cells in the induction of cRCS-X CFU, suggesting that LN cells contribute IL5. The level of CFU induced by an optimal dose of BCGF was enhanced further in the presence of LN cells and IL1.(ABSTRACT TRUNCATED AT 400 WORDS)
Physiological ecology of harmful algal blooms (1998)

Monoallelic expression (MAE) or extreme allele bias can account for incomplete penetrance, missing heritability and non-Mendelian diseases. In cancer, MAE is associated with shorter patient survival times and higher tumor grade. Prior studies showed that stochastic MAE is caused by stochastic epigenetic silencing, in a gene and tissue-specific manner. Here, we used C. elegans to study stochastic MAE in vivo. We found allele bias/MAE to be widespread within C. elegans tissues, presenting as a continuum from fully biallelic to MAE. We discovered that the presence of introns within alleles robustly decreases MAE. We determined that introns control MAE at distinct loci, in distinct cell types, with distinct promoters, and within distinct coding sequences, using a 5'-intron position-dependent mechanism. Bioinformatic analysis showed human intronless genes are significantly enriched for MAE. Our experimental evidence demonstrates a role for introns in regulating MAE, possibly explaining why some mutations within introns result in disease.
Peptidoglycan Hydrolase Fusions Maintain Their Parental Specificities (2006)

The increased incidence of bacterial antibiotic resistance has led to a renewed search for novel antimicrobials. Avoiding the use of broad-range antimicrobials through the use of specific peptidoglycan hydrolases (endolysins) might reduce the incidence of antibiotic-resistant pathogens worldwide. Staphylococcus aureus and Streptococcus agalactiae are human pathogens and also cause mastitis in dairy cattle. The ultimate goal of this work is to create transgenic cattle that are resistant to mastitis through the expression of an antimicrobial protein(s) in their milk. Toward this end, two novel antimicrobials were produced. The (i) full-length and (ii) 182-amino-acid, C-terminally truncated S. agalactiae bacteriophage B30 endolysins were fused to the mature lysostaphin protein of Staphylococcus simulans. Both fusions display lytic specificity for streptococcal pathogens and S. aureus. The full lytic ability of the truncated B30 protein also suggests that the SH3b domain at the C terminus is dispensable. The fusions are active in a milk-like environment. They are also active against some lactic acid bacteria used to make cheese and yogurt, but their lytic activity is destroyed by pasteurization (63 degrees C for 30 min). Immunohistochemical studies indicated that the fusion proteins can be expressed in cultured mammalian cells with no obvious deleterious effects on the cells, making it a strong candidate for use in future transgenic mice and cattle. Since the fusion peptidoglycan hydrolase also kills multiple human pathogens, it also may prove useful as a highly selective, multipathogen-targeting antimicrobial agent that could potentially reduce the use of broad-range antibiotics in fighting clinical infections.