Platelet PD-L1 suppresses anti-cancer immune cell activity in PD-L1 negative tumors.

Strategies that interfere with the binding of the receptor programmed cell death protein-1 (PD-1) to programmed death ligand-1 (PD-L1) have shown marked efficacy against many advanced cancers, including those that are negative for PD-L1. Precisely why patients with PD-L1 negative tumors respond to PD-1/PD-L1 checkpoint inhibition remains unclear. Here, we show that platelet-derived PD-L1 regulates the growth of PD-L1 negative tumors and that interference with platelet binding to PD-L1 negative cancer cells promotes T cell-induced cancer cytotoxicity. These results suggest that the successful outcomes of PD-L1 based therapies in patients with PD-L1 negative tumors may be explained, in part, by the presence of intra-tumoral platelets. Altogether, our findings demonstrate the impact of non-cancer/non-immune cell sources of PD-L1 in the tumor microenvironment in the promotion of cancer cell immune evasion. Our study also provides a compelling rationale for future testing of PD-L1 checkpoint inhibitor therapies in combination with antiplatelet agents, in patients with PD-L1 negative tumors.

Lactose drives Enterococcus expansion to promote graft-versus-host disease.

Disruption of intestinal microbial communities appears to underlie many human illnesses, but the mechanisms that promote this dysbiosis and its adverse consequences are poorly understood. In patients who received allogeneic hematopoietic cell transplantation (allo-HCT), we describe a high incidence of enterococcal expansion, which was associated with graft-versus-host disease (GVHD) and mortality. We found that Enterococcus also expands in the mouse gastrointestinal tract after allo-HCT and exacerbates disease severity in gnotobiotic models. Enterococcus growth is dependent on the disaccharide lactose, and dietary lactose depletion attenuates Enterococcus outgrowth and reduces the severity of GVHD in mice. Allo-HCT patients carrying lactose-nonabsorber genotypes showed compromised clearance of postantibiotic Enterococcus domination. We report lactose as a common nutrient that drives expansion of a commensal bacterium that exacerbates an intestinal and systemic inflammatory disease.

Impact of delayed insemination on pregnancy rates to gender selected semen in a fixed-time AI system.

The objectives of the current experiment were to determine if delaying insemination by 8 h in a FTAI protocol would alter estrus expression and pregnancy rates in cows inseminated with sex-sorted semen, characterize bull variation in pregnancy rates to sex-sorted semen and examine the impact of repeated years of FTAI to sex-sorted semen on calving distribution. Over three breeding seasons, postpartum cows (n = 839) were estrous synchronized using the 5-day CO-Synch + CIDR system. Cows were given GnRH (100 µg i.m., Factrel) at time of insertion of a controlled internal drug releasing device (CIDR; Eazi-Breed CIDR). Five d later CIDR was removed and PGF2α (25 mg i.m., Lutalyse) was given at removal and 8 h later. Estrus detection aids were applied at CIDR removal. Cows were inseminated with X-sorted or Y-sorted sex-sorted semen at 72 h (NORM) or 80 h (DELAY) after CIDR removal, and GnRH was administered at AI. At insemination, estrus status was categorized as positive (YES), partial (QUES), unknown (NR) or negative (NO). Bulls were introduced to cows at 14 d and removed at 60 d after FTAI. Pregnancy diagnosis was performed by ultrasound at d 60 after FTAI and via palpation at 60 d after bull removal. There was no difference (P > 0.05) in pregnancy rates to sex-sorted semen or final pregnancy rates between NORM and DELAY cows. Pregnancy to sex-sorted semen averaged 35.2% whereas final pregnancy rates were 90.6%. More cows (P < 0.05) in the DELAY group expressed estrus before FTAI, but this increase did not alter pregnancy rates to sex-sorted semen. Expression of estrus before FTAI increased (P < 0.02) pregnancy rates to sex-sorted semen across treatments with differences being YES > QUES or NR > NO. There was considerable variation in pregnancy rate by bull (P < 0.05) with pregnancy rates ranging from 55.6% to 19.3%. Whole herd calving distribution was altered (P < 0.05) after 3 y of use of sex-sorted semen compared to the previous 3 y when conventional semen was used. We conclude that delaying insemination by 8 h in an FTAI protocol did not improve pregnancy rates to sex-sorted semen despite more cows exhibiting estrus before FTAI. In addition, a high bull to bull variation in pregnancy rates to sex-sorted semen is a limitation in FTAI systems. Further research into FTAI strategies for use with sex-sorted semen is warranted.

Can liquorice extract and herbal solution prevent colonic mucosa damage caused by robenacoxib in dogs?

UNLABELLED: Non-steroidal anti-inflammatory drugs (NSAIDs) are commonly used in animals, especially in dogs, to manage pain due to inflammatory disease. This study investigated whether plant drugs can prevent mucosal injury induced by robenacoxib. We used fifteen healthy beagle dogs (7 male and 8 female) aged 4 months, weighing 4.2-5.1 kg at the beginning of the study. Endoscopy and biopsy of the colon were performed before and on the 21 day treatment with robenacoxib (1), robenacoxib, herbal solution with liquorice extract (2), placebo - an empty capsule (3). There were 5 animals in each group. The greatest microscopic damage in the colon was observed in animals which received robenacoxib. Plant drug administration reduced the severity of lesions in the colon when administered with robenacoxib (ARI = - 0.15). CONCLUSION: concurrent administration of liquorice extract and plant solution with robenacoxib was associated with significant decreased severity of the robenacoxib-induced colonic mucosal lesions.

Impact of pesticide contamination on aquatic microorganism populations in the littoral zone.

The effect of pesticide contamination of the littoral zone on the population of bacteria and fungi was analyzed using the example of a eutrophic water reservoir exposed for >30 years to the influence of expired crop-protection chemicals, mainly DDT. For three consecutive years, quantity analyses of bacteria and fungi were conducted and the composition of the microorganism population analyzed against seasonal dynamics. Mold and yeast-like fungi were also isolated and identified. Within the Bacteria domain, in addition to the large groups of microorganisms (Alphaprotobacteria, Betaprobacteria, and Gammaproteobacteria, Actinobacteria, and Cytophaga-Flavobacterium), the analysis also involved the presence of bacteria predisposed to degraded pesticides in natural environments: Pseudomonas spp. and Alcaligenes spp. The quantity dynamics of aquatic microorganisms indicated that bacteria and fungi under the influence of long-term exposure to DDT can adapt to the presence of this pesticide in water. No modifying effect of DDT was observed on the quantity of microorganisms or the pattern of seasonal relationships in the eutrophic lake. Changes were shown in the percentage share of large groups of bacteria in the community of microorganisms as was an effect of contamination on the species diversity of fungi. The data show the effectiveness of aquatic microorganism-community analyses as a tool for indicating changes in the water environment caused by pesticide contamination.

Investigation of the electronic, photosubstitution, redox, and surface properties of new ruthenium(II)-containing amphiphiles.

A series of pyridine- and phenol-based ruthenium(II)-containing amphiphiles with bidentate ligands of the following types are reported: [(L(PyI))Ru(II)(bpy)(2)](PF(6))(2) (1), [(L(PyA))Ru(II)(bpy)(2)](PF(6))(2) (2), [(L(PhBuI))Ru(II)(bpy)(2)](PF(6)) (3), and [(L(PhClI))Ru(II)(bpy)(2)](PF(6)) (4). Species 1 and 2 are obtained by treatment of [Ru(bpy)(2)Cl(2)] with the ligands L(PyI) (N-(pyridine-2-ylmethylene)octadecan-1-amine) and L(PyA) (N-(pyridine-2-ylmethyl)octadecan-1-amine). The imine species 3 and 4 are synthesized by reaction of [Ru(bpy)(2)(CF(3)SO(3))(2)] with the amine ligands HL(PhBuA) (2,4-di-tert-butyl-6-((octadecylamino)methyl)phenol), and HL(PhClA) (2,4-dichloro-6-((octadecylamino)methyl)phenol). Compounds 1-4 are characterized by means of electrospray ionization (ESI(+)) mass spectrometry, elemental analyses, as well as electrochemical methods, infrared and UV-visible absorption and emission spectroscopies. The cyclic voltammograms (CVs) of 1-2 are marked by two successive processes around -1.78 and -2.27 V versus Fc(+)/Fc attributed to bipyridine reduction. A further ligand-centered reductive process is seen for 1. The Ru(II)/Ru(III) couple appears at 0.93 V versus Fc(+)/Fc. The phenolato-containing 3 and 4 species present relatively lower reduction potentials and more reversible redox behavior, along with Ru(II/III) and phenolate/phenoxyl oxidations. The interpretation of observed redox behavior is supported by density functional theory (DFT) calculations. Complexes 1-4 are surface-active as characterized by compression isotherms and Brewster angle microscopy. Species 1 and 2 show collapse pressures of about 29-32 mN·m(-1), and are strong candidates for the formation of redox-responsive monolayer films.

The effect of isotonic multiple electrolyte infusions during anesthesia on blood gas and enzymatic values in horses.

General equine anesthesia is accompanied by the risk of complications and even death. The majority of general anesthetics used in veterinary practice lead to arterial hypotension, decreased capillary perfusion and ventilatory depression leading to pathological changes referred to as postanesthetic myopathy. In this study an attempt was made to analyze changes in the perioperative and postoperative acid-base equilibrium and blood enzymatic activity in horses, subject to the intensity of the applied fluid therapy. The horses were divided into two groups: I--administered Ringer's solution during surgery at a dose of 6-10 ml/kg BW/h, II--Ringer's solution administered at a dose of 12-15 ml/kg BW/h. The results of the study indicate that use of hydrating fluid infusions during general anesthesia, including infusions without hypertensive substances, minimizes the risk of myocyte damage due to disturbances in local and general blood circulation.

Selected fluorescent techniques for identification of the physiological state of individual water and soil bacterial cells - review.

Stimulated by demands of the natural environment conservation, the need for thorough structural and functional identification of microorganisms colonizing different ecosystems has contributed to an intensive advance in research techniques. The article shows that some of these techniques are also a convenient tool for determination of the physiological state of single cells in a community of microorganisms. The paper presents selected fluorescent techniques, which are used in research on soil, water and sediment microorganisms. It covers the usability of determination of the dehydrogenase activity of an individual bacterial cell (CTC+) and of bacteria with intact, functioning cytoplasmic membranes, bacteria with an integrated nucleiod (NuCC+) as well as fluorescent in situ hybridization (FISH).

On contemporaneous controls, unlikely outcomes, boxes and replacing the 'Student': good statistical practice in pharmacology, problem 3.

This paper is intended to assist pharmacologists to make the most of statistical analysis and avoid common errors. A scenario, in which an experimenter performed an experiment in two separate stages, combined the control groups for analysis and found some surprising results, is presented. The consequences of combined controls are discussed, appropriate display and analysis of the data are described, and an analysis of the likelihood of erroneous conclusions is made. Comparisons between data from separately conducted experimental series are hazardous when there is any possibility that the properties of the experimental units have changed between the series. Experiments that have been performed independently should be analyzed independently. Unlikely or surprising results should be treated with caution and a high standard of evidence should be required, and verification by repeated experiments should be performed and reported. Box and whisker plots contain more information than plots more commonly used to display for qualitative variables and should be used where the sample size is large enough (say, n > or = 5). In most biomedical experiments the observations are not random samples from large populations as assumed by conventional parametric analyses such as Student's t-test, and so permutation tests, which do not lose their validity when a sampled population is non-normal or when the data are not random samples, should frequently be used instead of Student's t-tests.

Haplotypes and gene expression implicate the MAPT region for Parkinson disease: the GenePD Study.

BACKGROUND: Microtubule-associated protein tau (MAPT) has been associated with several neurodegenerative disorders including forms of parkinsonism and Parkinson disease (PD). We evaluated the association of the MAPT region with PD in a large cohort of familial PD cases recruited by the GenePD Study. In addition, postmortem brain samples from patients with PD and neurologically normal controls were used to evaluate whether the expression of the 3-repeat and 4-repeat isoforms of MAPT, and neighboring genes Saitohin (STH) and KIAA1267, are altered in PD cerebellum. METHODS: Twenty-one single-nucleotide polymorphisms (SNPs) in the region of MAPT on chromosome 17q21 were genotyped in the GenePD Study. Single SNPs and haplotypes, including the H1 haplotype, were evaluated for association to PD. Relative quantification of gene expression was performed using real-time RT-PCR. RESULTS: After adjusting for multiple comparisons, SNP rs1800547 was significantly associated with PD affection. While the H1 haplotype was associated with a significantly increased risk for PD, a novel H1 subhaplotype was identified that predicted a greater increased risk for PD. The expression of 4-repeat MAPT, STH, and KIAA1267 was significantly increased in PD brains relative to controls. No difference in expression was observed for 3-repeat MAPT. CONCLUSIONS: This study supports a role for MAPT in the pathogenesis of familial and idiopathic Parkinson disease (PD). Interestingly, the results of the gene expression studies suggest that other genes in the vicinity of MAPT, specifically STH and KIAA1267, may also have a role in PD and suggest complex effects for the genes in this region on PD risk.

Continued challenges with the use of erythropoiesis-stimulating agents in patients with cancer: perspectives and issues on policy-guided health care.

Erythropoiesis-stimulating agents (ESAs) are approved as an alternative to blood transfusions for treating anemia secondary to chemotherapy in patients with cancer. Recently, ESAs have been a source of controversy and confusion in the oncology community. This began when two European trials-the Breast Cancer Erythropoietin Survival Trial (BEST) and the Advanced Head-and-Neck Cancer Treated with Radiotherapy (ENHANCE) Study-raised safety concerns about decreased overall survival and increased venous thromboembolic events. In 2004, the United States Food and Drug Administration (FDA) convened its Oncologic Drugs Advisory Committee (ODAC) to review the data and reassess the risks and benefits of ESAs in patients with cancer. On May 10, 2007, ODAC reconvened when five trials (BEST, ENHANCE, AMG-20010103, AMG-20000161, and EPO-CAN-20) showed decreased overall survival. The briefing document noted that studies demonstrating detrimental effects on survival and/or tumor outcomes used an unapproved treatment regimen designed to maintain hemoglobin levels above 12 g/dl. On May 14, 2007, just days after the ODAC reconvened, the Centers for Medicare and Medicaid Services (CMS) released a proposed decision memo for a national coverage determination (NCD) imposing restrictions on ESAs. For health care providers, aspects of the proposed NCD were markedly inconsistent with FDA-approved ESA use and generally were considered ambiguous and unclear. Over objections of several professional associations and members of Congress, on July 30, 2007, CMS posted the final NCD and declared it effective immediately. When compared with FDA-approved labeling and professional society guidelines, the NCD revealed differences in ESA initiation, dosage escalation, dosage reduction, and definition of response. These discrepancies have generated confusion among health care providers, who are struggling over whether they can feasibly provide a dual system of care-one for Medicare patients and another for non-Medicare patients-that is evidence based. With this supplement, we hope to educate health care providers on the issues and challenges associated with policy-guided health care when discrepancies exist between the policy and evidence-based practice; offer guidance on implementing the NCD; and highlight the important role of pharmacists in the process.

Cyclodialysis in the treatment of nonhealing secondary glaucoma.

In cases of glaucoma with elevated intra ocular pressure, the first objective of treatment is its reduction and normalization. In cases when excessive pressure has resulted in a loss of vision, the aim of treatment is to attenuate the pain and to improve the comfort of the patient. This paper presents a case of glaucoma secondary to anterior uveitis, resistant to pharmacological treatment. A gonioscopic examination in both eyes revealed a narrowed filtration angle. An ophthalmoscopic examination of the fundus showed partial retinal detachment. The cyclodialysis procedure was performed in both eyes. In the post-surgery period, intraocular pressure (IOP) in both eyes showed a downward tendency. The treatment of glaucoma secondary to anterior uveitis, particularly in its advanced state, often requires the combination of pharmacological treatment with a surgical procedure. Positive results in maintaining the patency of the fistula were observed after the application of a steroid therapy and a regular massage of filtration bleb, which significantly contributed to maintaining IOP at a desired level.

Short-term results of Ahmed glaucoma valve implantation in the surgical treatment of angle-recession glaucoma in dog.

Angle-recession glaucoma, being a consequence of blunt injury of an eyeball, is likely to develop in both a short or very distant period. The likelihood of the incidence of that condition in the clinical form is determined by the extent of injuries of ciliary cleft structures, especially of a trabecular meshwork. The manuscript reports a case of an early glaucoma secondary to a blunt injury of an eyeball with symptoms of angle recession to 300 degrees of periphery and intraocular pressure (IOP) at a level of 62 mmHg. The basic diagnostic examination that enables the evaluation of an angle, iris root and an opening to the ciliary cleft is gonioscopy. In case of a lack of response to the pharmacological reduction of IOP, a method of choice still remains a surgical procedure. In the reported case, a drainage procedure was applied in the form of implantation of the Ahmed valve prosthesis. Control tonometric examinations of the operated eye, performed over a 20-week period of postoperative management, demonstrated IOP at a level of 19 mmHg.

Good statistical practice in pharmacology. Problem 2.

BACKGROUND AND PURPOSE: This paper is intended to assist pharmacologists to make the most of statistical analysis and in avoid common errors. APPROACH: A scenario is presented where an experimenter performed an experiment to test the effects of two drugs on cultured cells. Analysis of the results, expressed as percentage of control, by a one-way ANOVA yielded P=0.058 and the experimenter concluded that neither drug was effective. The data were expressed as percentage of control because of pairing of the data within each experimental run, a common feature in cell culture experiments. Such data can be analysed with potentially more powerful ANOVA methods equivalent to the paired t-test. Monte Carlo simulations are presented to compare the power of relevant analyses. RESULTS: For data correlated within experimental run (i.e. paired values), transformation to percentage of control improved the power of a one-way ANOVA to detect a real effect, but a randomized block ANOVA (equivalent to a 2-way ANOVA with experiment and treatment as factors) using the raw values was substantially more powerful. The randomized block ANOVA performed well even with uncorrelated data, being only marginally less powerful than the one-way ANOVA. CONCLUSIONS AND IMPLICATIONS: A randomized block ANOVA is far superior to the one-way ANOVA with correlated data, and with uncorrelated data it is only marginally less powerful. Thus where there is, or might reasonably be, such a correlation (e.g. relatedness among the data within a single experimental run, or within a multi-well culture plate, or within an animal, et cetera), use the more powerful randomized block ANOVA rather than one-way ANOVA.

Good statistical practice in pharmacology. Problem 1.

BACKGROUND AND PURPOSE: This paper is intended to assist pharmacologists in making the most of statistical analysis and in avoiding common errors that can lead to false conclusions. APPROACH: A scenario is presented where a pathway inhibitor increased blood pressure responses to an agonist by about one third. The fictional experimenter concludes that the inhibitor enhanced the responses to the agonist, but has not applied any statistical analysis. Questions are asked of the reader, and a discussion of the author's answers is presented. RESULTS: The agonist responses have unequal standard errors, as often seen in data like these concentration-response curves with responses expressed as change from baseline. The uneven variability (heteroscedasticity) violates an assumption of conventional parametric statistical analyses, but can be corrected by data transformation. Expressing the data as absolute blood pressure and then transforming it to log blood pressure eliminated the heteroscedasticity, but made evident an effect of the inhibitor on baseline blood pressure. CONCLUSIONS AND IMPLICATIONS: Statistical analysis is a sensible precaution against mistakes, but cannot protect against all erroneous conclusions. In this scenario, the inhibitor reduced the blood pressure and increased responses to the agonist. However, it is likely that the latter effect was a consequence of the former and thus no conclusion can be safely drawn about any direct interaction between the agonist and the pathway inhibitor from this experiment. Where results are awkward to interpret because of confounding factors such as an altered baseline, statistical analysis may not be very useful in supporting a safe conclusion.

Herbicide exposure modifies GSTP1 haplotype association to Parkinson onset age: the GenePD Study.

BACKGROUND: Polymorphisms in the glutathione S-transferase pi gene (GSTP1), encoding GSTP1-1, a detoxification enzyme, may increase the risk of Parkinson disease (PD) with exposure to pesticides. Using the GenePD Study sample of familial PD cases, we explored whether GSTP1 polymorphisms were associated with the age at onset of PD symptoms and whether that relation was modified by exposure to herbicides. METHODS: Seven single-nucleotide polymorphisms (SNPs) were genotyped and tested for association with PD onset age in men in three strata: no exposure to herbicides, residential exposure to herbicides, and occupational exposure to herbicides. Haplotypes were similarly evaluated in stratified analyses. RESULTS: Three SNPs were associated with PD onset age in the group of men occupationally exposed to herbicides. Three additional SNPs had significant trends for the association of PD onset age across the herbicide exposure groups. Haplotype results also provided evidence that the relation between GSTP1 and onset age is modified by herbicide exposure. One haplotype was associated with an approximately 8-years-earlier onset in the occupationally exposed group and a 2.8-years-later onset in the nonexposed group. CONCLUSIONS: Herbicide exposure may be an effect modifier of the relation between glutathione S-transferase pi gene polymorphisms and onset age in familial PD.

Structural and electronic behavior of unprecedented five-coordinate iron(III) and gallium(III) complexes with a new phenol-rich electroactive ligand.

A new asymmetric pentadentate ligand was designed to impose low symmetry to trivalent ions. Five-coordinate Fe3+ and Ga3+ complexes were investigated by crystallographic, electrochemical, and electron paramagnetic resonance methods showing enhanced redox reversibility. Calculations were performed to account for the observed trends.

BDNF genetic variants are associated with onset age of familial Parkinson disease: GenePD Study.

Brain-derived neurotrophic factor (BDNF) stimulates neuronal growth and protects nigral dopamine neurons in animal models of Parkinson disease (PD). Therefore, BDNF is a candidate gene for PD. The authors investigated five single-nucleotide polymorphisms in 597 cases of familial PD. Homozygosity for the rare allele of the functional BDNF G196A (Val66Met) variant was associated with a 5.3-year older onset age (p = 0.0001). These findings suggest that BDNF may influence PD onset age.

Influence of ligand rigidity and ring substitution on the structural and electronic behavior of trivalent iron and gallium complexes with asymmetric tridentate ligands.

Species 1-6 are [M(III)(L)2]ClO4 complexes formed with the PhO--CH=N-CH2-Py imines, (L(I))- and (L(tBuI))-, and PhO--CH2-NH-CH2-Py amines, (L(A))- and (L(tBuA))-, in which PhO- is a phenolate ring and Py is a pyridine ring and the prefix tBu indicates the presence of tertiary butyl groups occupying the positions 4 and 6 of the phenol ring. Monometallic species with d5 high-spin iron (1, 2, 3, 4) and d10 gallium (5, 6) were synthesized and characterized to assess the influence of the ligand rigidity and the presence of tertiary butyl-substituted phenol rings on their steric, electronic, and redox behavior. Characterization by elemental analysis, mass spectrometry, IR, UV-visible, and EPR spectroscopies, and electrochemistry has been performed, and complexes [FeIII(L(tBuI))2]ClO4 (2), [FeIII(L(tBuA))2]ClO4 (4), and [Ga(III)(L(tBuI))2]ClO4 (5) have been characterized by X-ray crystallography. The crystal structures show the imine ligands meridionally coordinated to the metal centers, whereas the amine ligands are coordinate in a facial mode. Cyclic voltammetry shows that the complexes with the ligands (L(tBuI))- and (L(tBuA))- were able to generate ligand-based phenoxyl radicals, whereas unsubstituted ligands displayed ill-defined redox processes. EPR spectroscopy supports high-spin configurations for the iron complexes. UV-visible spectra are dominated by charge-transfer phenomena, and imine compounds exhibit dramatic hyperchromism when compared to equivalent amines. The tertiary butyl groups on the phenolate ring enhance this trend. Detailed B3LYP/6-31G(d)-level calculations have been used to account for the results observed.

Laparoscopic cryptorchidectomy in dogs --report of 15 cases.

The study aimed at determining the usability of laparoscopic cryptorchidectomy in treating cases of simple cryptorchidism and neoplastic testes in dogs. The presence of one or both testes inside the peritoneal cavity was confirmed in this study by an ultrasonographic examination employing the use of a 6.5 MHz convex head. The surgeries were conducted on a group of 15 dogs. In 12 subjects a one-sided cryptorchidism was found, 9 of which had right-sided cryptorchidism. In 3 dogs, both-sided cryptorchidism was observed. In 3 cases, an increase in the size of the abdominal testes with the symptoms of hyperestrogenism and feminisation raised a suspicion of neoplastic changes (SCT) taking place, which was confirmed by a histopathological examination. A laparoscopic cryptorchidectomy with intracorporal ligation was performed in all the subjects. The spermatic cord and ductus deferens were occluded with a ligature of an absorbable material, and a manually applied knot.