R1 pancreatic cancer patients benefit from chemoradiation when initiation of adjuvant therapy is delayed.

BACKGROUND: Adjuvant chemotherapy (AC) following pancreaticoduodenectomy (PD) for pancreas cancer (PDAC) has been demonstrated to improve survival. However, the optimal adjuvant treatment (AT) regimen for R1-margin patients remains unclear. This retrospective study investigates the impact of AC vs. adjuvant chemoradiotherapy (ACRT) on survival (OS). MATERIAL AND METHODS: The NCDB was queried for patients with PDAC who underwent PD between 2010 and 2018. Patients were divided into, (A) AC<60 days, (B) ACRT<60 days, (C) AC≥60 days, and (D) ACRT≥60 days. Kaplan-Meier survival analyses and Cox multivariable regression analyses were performed. RESULTS: Among 13 740 patients, median OS was 23.7 months. For R1 patients, median OS for timely AC and ACRT, and delayed AC and ACRT was 19.91, 19.19, 15.24, 18.96 months, respectively. While time of AC initiation was an insignificant factor for R0 patients (p = 0.263, CI 0.957-1.173), a survival benefit was found for R1 patients who received AC<60 vs. ≥60 days (p = 0.041, CI 1.002-1.42). Among R1 patients, administration of delayed ACRT achieves the same survival benefit of timely AC initiation (p = 0.074, CI 0.703-1.077). CONCLUSION: The study suggests value in ACRT for patients with R1 margins when delay of AT≥60 days cannot be avoided. Hence, ACRT may mitigate the negative impact of delayed AT initiation for R1-patients.

A one-health perspective: use of hemoderivative regenerative therapies in canine and equine patients.

Regenerative medicine therapies have become significant tools for treatment of joint, soft tissue, and a variety of other conditions in animals and humans. Regenerative medicine aims to restore form and function of injured tissues using the body's own resources such as cells, fluids (ie, plasma and serum), and their resulting anti-inflammatory and prohealing cytokines. Platelet-rich plasma and other hemoderivatives have application for joint disorders such as osteoarthritis, cartilage injury, synovitis, and soft tissue injuries. These therapies achieve anti-inflammatory and healing effects without the use of corticosteroid therapy. This response is an advantage when treating young animals or human patients, and in animals with metabolic or hormonal issues such as equine pituitary pars intermedia dysfunction. Also, these therapies may have beneficial effects when traditional IA treatments such as corticosteroids and/or hyaluronan are no longer effective at reducing joint inflammation and pain. Examples of hemoderivative regenerative therapies to be discussed include platelet-rich plasma, autologous conditioned serum, autologous protein solution, and α-2 macroglobulin.

Practical Rehabilitation and Physical Therapy for the General Equine Practitioner.

Physical treatment and rehabilitation play major roles in recovery and maintenance of the equine athlete, and many therapeutic measures are accessible by the veterinarian in general practice. An accurate diagnosis of the condition undergoing treatment is a requirement, and measurable parameters obtained at diagnosis allows for quantification of treatment outcomes. Therapeutic modalities accessible to the general practicing veterinarian are reviewed. Mechanisms of action, indications, and treatment protocols of thermal therapy, therapeutic ultrasound, extracorporeal shock wave, and laser are discussed. Manipulative therapies, including stretching and use of core strengthening exercises and equipment, are outlined.

Binding inhibitors of the bacterial sliding clamp by design.

The bacterial replisome is a target for the development of new antibiotics to combat drug resistant strains. The ß(2) sliding clamp is an essential component of the replicative machinery, providing a platform for recruitment and function of other replisomal components and ensuring polymerase processivity during DNA replication and repair. A single binding region of the clamp is utilized by its binding partners, which all contain conserved binding motifs. The C-terminal Leu and Phe residues of these motifs are integral to the binding interaction. We acquired three-dimensional structural information on the binding site in ß(2) by a study of the binding of modified peptides. Development of a three-dimensional pharmacophore based on the C-terminal dipeptide of the motif enabled identification of compounds that on further development inhibited α-ß(2) interaction at low micromolar concentrations. We report the crystal structure of the complex containing one of these inhibitors, a biphenyl oxime, bound to ß(2), as a starting point for further inhibitor design.

Synthesis, binding and bioactivity of gamma-methylene gamma-lactam ecdysone receptor ligands: advantages of QSAR models for flexible receptors.

Nuclear hormone receptors, such as the ecdysone receptor, often display a large amount of induced fit to ligands. The size and shape of the binding pocket in the EcR subunit changes markedly on ligand binding, making modelling methods such as docking extremely challenging. It is, however, possible to generate excellent 3D QSAR models for a given type of ligand, suggesting that the receptor adopts a relatively restricted number of binding site configurations or 'attractors'. We describe the synthesis, in vitro binding and selected in vivo toxicity data for gamma-methylene gamma-lactams, a new class of high-affinity ligands for ecdysone receptors from Bovicola ovis (Phthiraptera) and Lucilia cuprina (Diptera). The results of a 3D QSAR study of the binding of methylene lactams to recombinant ecdysone receptor protein suggest that this class of ligands is indeed recognised by a single conformation of the EcR binding pocket.

Parasiticidal 2-alkoxy- and 2-aryloxyiminoalkyl trifluoromethanesulfonanilides.

A series of novel 2-alkoxy- and 2-aryloxyiminoalkyl trifluoromethanesulfonanilide derivatives have shown significant in vitro parasiticidal activity against the ectoparasites Ctenocephalides felis and Rhipicephalus sanguineus. A number of these compounds also displayed significant in vitro endoparasite activity against the nematode Haemonchus contortus.

Ring expansion reactions of 4-amino-1,1-dioxo-[1,2,3,5]-thiatriazoles.

An unusual ring-expansion reaction of 4-amino-1,1-dioxo-[1,2,3,5]-thiatriazoles has been identified that produces the relatively rare 5-amino-1,1-dioxo-[1,2,4,6]-thiatriazines and. Initial alkylation of the thiatriazole with alpha-halo-esters at N-3 produces alpha-substituted esters which, under basic reaction conditions, undergo opening of the thiatriazole ring and re-closure to a thiatriazine ring. Similar alkylations of with diethyl chloromalonate and ethyl dichloroacetate lead to the loss of SO2 and the production of triazine and triazole, apparently by an initial alkylation at N-5. The reaction of with phenacyl bromides or a phenacyl dibromide forms fully unsaturated 5-amino-1,1-dioxo-[1,2,4,6]-thiatriazines.

Comparison of hematologic values and transforming growth factor-beta and insulin-like growth factor concentrations in platelet concentrates obtained by use of buffy coat and apheresis methods from equine blood.

OBJECTIVE: To evaluate the buffy coat and apheresis methods for preparation of platelet concentrates from equine blood by comparing platelet and growth factor concentrations. ANIMALS: 15 mature mixed-breed geldings. PROCEDURE: Whole blood samples were collected and processed by use of a buffy coat or apheresis method to obtain platelet poor and platelet concentrated fractions. The PCV, WBC count, and platelet count were compared among whole blood samples, platelet poor fractions, concentrates obtained by use of the apheresis method (ie, apheresis platelet concentrates), and concentrates obtained by use of the buffy coat method (ie, buffy coat platelet concentrates). Concentrations of transforming growth factor-beta (ie,TGF-beta1 andTGF-beta2) and insulin-like growth factor were compared between buffy coat and apheresis platelet concentrates. RESULTS: Platelet concentrations were 8.9-fold and 5.2-fold greater in buffy coat and apheresis platelet concentrates, respectively, compared with whole blood. Platelet concentrations were 13.1-fold greater in filtered apheresis platelet concentrates, compared with whole blood. TGF-beta1 concentrations were 2.8-fold and 3.1-fold greater in buffy coat and apheresis platelet concentrates, respectively, and TGF-beta1 concentrations were 10.5-fold greater in filtered apheresis platelet concentrates, compared with whole blood. TGF-beta2 concentrations were 3.6-fold greater in apheresis platelet concentrates, compared with whole blood. Platelet concentrations correlated with growth factor concentrations across all blood and platelet fractions. White blood cell counts had a significant positive correlation with TGF-beta1 concentration in buffy coat platelet concentrates. CONCLUSIONS AND CLINICAL RELEVANCE: Platelets and TGF-beta1 can be concentrated reliably from equine blood by use of buffy coat or apheresis methods, without modification of the protocols used for humans.

Lontophoretic administration of dexamethasone into the tarsocrural joint in horses.

OBJECTIVE: To determine whether iontophoretic administration of dexamethasone to horses results in detectable concentrations in synovial fluid, plasma, and urine. ANIMALS: 6 adult mares. PROCEDURE: Iontophoresis was used to administer dexamethasone. Treatments (4 mA for 20 minutes) were administered to a tarsocrural joint of each mare. The drug electrode contained 3 ml of dexamethasone sodium phosphate at a concentration of 4 or 10 mg/ml. Samples of synovial fluid, blood, and urine were obtained before and 0.5, 4, 8, and 24 hours after each treatment. All samples were tested for dexamethasone using an ELISA. Synovial fluid also was evaluated for dexamethasone, using high-performance liquid chromatography. RESULTS: The lower and upper limits of detection for dexamethasone in synovial fluid with the ELISA were 0.21 and 1.5 ng/ml, respectively. Dexamethasone administered at a concentration of 10 mg/ml was detected by the ELISA in synovial fluid of 5 mares from 0.5 to 24 hours and in urine of 4 mares from 0.5 to 8 hours after each treatment, but it was not detected in plasma. Mean synovial fluid concentration of dexamethasone was 1.01 ng/ml. Dexamethasone administered at a concentration of 4 mg/ml was detected by the ELISA in urine of 2 mares at 0.5 and 4 hours after treatment, but it was not detected in synovial fluid or plasma. CONCLUSIONS AND CLINICAL RELEVANCE: Iontophoresis cannot be considered an effective method for delivery of dexamethasone to synovial fluid of horses, because drug concentrations achieved in this study were less than therapeutic concentrations.

Characterization of the three immunoglobulin G subclasses of macaques.

Southern blot experiments with genomic DNA samples of rhesus monkeys and crab-eating macaques and human C gamma-specific probes indicated that the two macaque species studied here possessed three C gamma genes per haploid genome. By amplifying the cDNA from macaque-mouse hybridomas, the coding sequences of two different rhesus monkey immunoglobulin (Ig)G subclasses, IgG1rh (Cgamma1rh) and IgG2rh (Cgamma2rh), and one crab-eating macaque IgG subclass IgG1mafa (Cgamma1mafa), were characterized. None of the 16 rhesus monkey-mouse hybridomas studied here secreted IgG of the third subclass IgG3rh (Cgamma3rh). The Cgamma3rh gene was partly characterized at the genomic level. The cDNA of the Cgamma3rh gene was amplified from mRNA of rhesus monkey peripheral blood mononuclear cells (PBMC). The results are analysed in terms of phylogenesis of the C gamma genes. The cDNA sequences coding for the Cmu and the Ckappa domains of rhesus monkey Ig were established and compared to their human and non-human primate counterparts.

Variable region gene segment utilization in rhesus monkey hybridomas producing human red blood cell-specific antibodies: predominance of the VH4 family but not VH4-21 (V4-34).

Structural analyses of human immunoglobulin gene segments from monoclonal cell lines provide valuable information regarding the antibody repertoire. This information, in conjunction with a nearly complete knowledge of the human immunoglobulin germline repertoire, now allows further investigation into the underlying molecular basis responsible for some of the observed biases found in the expressed repertoire. One human heavy chain variable region gene segment, V4-34 (VH4-21), is one of the most prevalent gene segments in the expressed repertoire. The overwhelming presence of the V4-34 gene segment suggests that it may play an important role in immune responses. However, there is currently little information regarding its presence and potential importance in nonhuman primates. In order to determine if this gene segment is used by lower primates in a similar manner we determined the molecular structure of the variable region gene segments that are expressed by macaque monoclonal heterohybridomas that are specific for human red blood cell antigens. Eleven of the 12 hybridomas are derived from Rhesus monkeys (Macaca mulatta) and one is from a cynomologous monkey (Macaca fascicularis), all of which have been immunized with human red blood cells. The predominance of a VH4-like family and the specific absence of a VH4-21 equivalent led us to further characterize the macaque VH4 gene family at the germline level. Therefore, germline gene segments from the macaque equivalent to the human VH4 gene family are also described.

The human antibody repertoire: heavy and light chain variable region gene usage in six alloantibodies specific for human HLA class I and class II alloantigens.

Peripheral blood B lymphocytes have been isolated from healthy individuals who were immunized with lymphocytes from HLA-incompatible donors and transformed with Epstein-Barr virus to produce human monoclonal cell lines specific for human HLA molecules. The cell lines have been previously characterized and are known to bind to various class I and class II alloantigens. In this report we describe the molecular characterization of the heavy and light chain variable region gene segments that are utilized by these monoclonal antibodies. Using the polymerase chain reaction and primer pairs specific for the respective constant region and VH or VL family, rearranged variable region gene segments were amplified from cDNA from individual cell lines. Products were then subcloned, sequenced and analysed for gene usage and apparent somatic mutation. The results show that the VH3 gene family predominates in a group of six heavy chains (four out of six) with one VH1 and one VH4 gene segment. The light chain variable region gene family usage is more diverse with 2 V kappa 3, 1 V kappa 1, 2 V lambda 2 and 1 V lambda 3. The extent of apparent somatic mutation is minimal, relative to our previous observations in a group of high affinity human monoclonal antibodies specific for pathogenic organisms.

Faculty authoring of course-specific software for disadvantaged nursing students using LinkWay: a case study.

This article adds to the literature on the use of hypermedia systems for software development in nursing education by describing design and authoring considerations in the construction of course-specific remediating software. In 1991, under the guidance of an instructional designer, faculty members developed LinkWay folders on aspects of the endocrine system for remedial use by students in Project Get Ahead in Nursing at Southern Illinois University at Edwardsville. Existing content outlines were modified according to design criteria, including representing information graphically, making the software translation interactive, embedding visual prompts and cues, and conforming to a specific design protocol. The use of LinkWay in facilitating the implementation of both procedural of both procedural and screen protocol is discussed, and several screen templates are presented, including menus, multiple choice, short answer, multiple choice browsing, and end-of-section evaluation. Current project status and conclusions for remediation using a hypermedia approach are discussed.

The molecular structure of human antibodies specific for the human immunodeficiency virus.

The molecular structure of human antibodies that are specific for human immunodeficiency virus-1 (HIV-1) are of increasing interest as AIDS research progresses toward passive immunotherapeutics in the maintenance and prevention of infection. In recent years a number of human, HIV-specific hybridomas and EBV-transformed B cell lines, as well as a combinatorial library, have been developed and characterized at the molecular level. These sources have provided valuable information on the immunoglobulin heavy- and light-chain variable-region gene usage and the extent and appearance of somatic mutation in a disease where the immune system is under constant stimulation over a long period of time. In this article we review the current data available on the molecular structure of these antibodies.

Molecular characterization of human antibodies to bacterial antigens: utilization of the less frequently expressed VH2 and VH6 heavy chain variable region gene families.

Structural analysis of the human immunoglobulin repertoire holds promise for determining the basis of variable region gene usage in response to a variety of auto and exogenous antigens. Here we report the nucleotide sequences of the heavy and light chain variable regions expressed by three human monoclonal antibodies specific for two clinically relevant bacterial pathogens, Bordetella pertussis and Haemophilus influenzae type b. The cell lines were derived by in vitro stimulation of lymphocytes from spleen or tonsillar tissue, respectively, and bind to different antigens from the two organisms. The single B. pertussis antibody is of the IgM lambda isotype and utilizes the single VH6 gene segment in combination with a V lambda 2 gene and demonstrates limited somatic mutation, yet is highly indicative of an antigen-driven immune response. One H. influenzae antibody is of the IgG2 lambda isotype and expresses a VH3 gene segment with a V lambda 1 gene, while the second cell line produces an IgG3 lambda antibody expressing a combination of VH2/V lambda 3. Both molecules show evidence of somatic mutation. The D gene segments of the heavy chains vary in length and display limited sequence homology with known germline D segments. As demonstrated previously, JH4 predominates (two JH4 and one JH3) and all three utilize the J lambda 3 gene segment. In addition, we have isolated and sequenced a number of germline VH2 gene segments in an attempt to better understand the nature of the VH2 germline repertoire. In addition to contributing to the understanding of the human antibody repertoire, such clinically relevant molecules may prove to be a source of passive immunotherapy for those at risk to developing disease.

Variable region genes of anti-HIV human monoclonal antibodies: non-restricted use of the V gene repertoire and extensive somatic mutation.

The extent of the expressed human V gene repertoire for the most part has been derived from fetal cDNA libraries, autoantibodies, and myeloma proteins. In order to continue to explore the utilization of the VH and VL gene repertoire in response to exogenous viral antigens, the heavy and light chain cDNAs from four human anti-HIV monoclonal antibodies were PCR amplified from human-mouse heterohybridomas, cloned, and nucleotide sequence analysis performed. Of the monoclonals analyzed, three were directed against gp120 and one reacted with gp41. Three of the antibodies were of the IgG1 lambda isotype and one was an IgG1 kappa. Three of the four heavy chains were derived from VHI gene segments and one VHII was observed. D segments showed evidence of D-D joining and three JH4 and one JH5 gene were utilized. Two V lambda II lambda chains and one from the V lambda III gene family were observed and the single kappa chain sequenced was from the V kappa III family. DNA sequence comparison with known germline gene segments identified putative precursor V gene segments for one of the heavy chains and two light chains. Comparison of the expressed amino acid sequences with the predicted germline sequences indicated that changes were clustered in the CDRs and FR3 regions of the V gene segments. We reported previously the nucleotide sequences of five human monoclonal antibodies from HIV-infected individuals, three of which utilized VHIV, one VHV and one a VHI gene segment and also found extensive evidence of somatic mutation. Collectively, our results indicate that an antigen driven response is functioning following HIV infection and, surprisingly, to date we have not encountered a VHIII gene segment. Since VHIII is the largest human VH gene family, it may well be that this under-representation has both functional and clinical implications.

Probing the human antibody repertoire to exogenous antigens. Characterization of the H and L chain V region gene segments from anti-hepatitis B virus antibodies.

Structural studies of human antibody V regions have been largely limited to those involving the fetal repertoire, autoantibodies, and malignant cell rearrangements, leaving the "normal" repertoire relatively unexplored. In this study we describe the nucleotide sequences of the H and L chain V regions of four antibodies specific for the surface Ag of the hepatitis B virus. Monoclonal cell lines were derived from healthy individuals who received standard immunizations with the serum-derived or recombinant hepatitis B virus vaccines by fusion of PBL to a heterohybridoma cell line, SPAZ-4. We utilized the polymerase chain reaction to amplify the H and L chain V regions for cloning and sequencing. The four antibodies express the following V region combinations: VHIII/V lambda V, VHIII/V kappa II, VHIV/V kappa I, VHV/V lambda III. When compared to germline genes with the closest sequence homology, all of the V regions appear to have undergone somatic mutation, ranging from 3.4 to 11.3% for the H chain, and 5.1 to 9.2% for the L chain. Analysis of the mutations shows them to be typical for an Ag-driven immune response.

Molecular characterization of five human anti-human immunodeficiency virus type 1 antibody heavy chains reveals extensive somatic mutation typical of an antigen-driven immune response.

We report the heavy chain variable region sequences from the cDNAs of five previously described monoclonal cell lines producing human antibodies specific for the human immunodeficiency virus type 1 and detail the molecular characteristics, germ-line origins, and extent of somatic mutation among these antibodies. Three of the five heavy chain variable regions derive from the VHIV gene family, but each has arisen from a different heavy chain variable region (VH) gene segment within the VHIV family. In addition, one is derived from a VHI gene segment, and one is derived from a VHV gene segment. Since four of the five antibodies arise from known germ-line VH elements, a precise determination of the extent of somatic variation is possible. The amount of variation from the closest germ-line sequence ranges from 4.5% to 14.8% among these antibodies, most of which is concentrated in the complementarity-determining regions. In general, the diversity (D) segments are long, characteristic of D-D fusions and/or extensive terminal deoxynucleotidyltransferase activity; however, definitive homologies cannot be found with the known germ-line D segments. Joining (JH) gene segment utilization appears random. The use of five different germ-line VH gene segments and extensive somatic mutation provides evidence that a polyclonal, antigen-driven immune response occurs during the natural infection with human immunodeficiency virus.