The impact of immunosuppression on development of UC its

The impact of immunosuppression on development of UC, its intensity, and the type of drugs has not been well investigated. More intensive immunosuppression and a combination of different types of agents have been associated with a higher incidence of UC. Furthermore, treatment with anti-T-lympocyte globulin (ATG) is associated with elevated risk of posttransplant lymphoproliferative disorders but the treatment\’s link to UC has not been investigated. Infection with BK and SV40 viruses may be an additional risk factor for development of UC in this instance.

In this one-center review, the authors make the following points: urothelial cancer is a relatively common genitourinary malignancy in kidney transplant recipients, risk factors should trigger screening and surveillance protocols for the disease in kidney transplant candidates and recipients, and treatments for urothelial cancers in this patient population need to be tailored for surgical anatomy and immunosuppression.
Age- and gender-adjusted standardized incidence ratios (SIRs) are useful to compare observed vs expected incidence rates of urothelial cancers after kidney transplantation. They have been reported to be 2.0 for Canada and 42.9 for Taiwan. The very high SIR for the latter is probably due to the urothelial and nephropathy risk factors of aristolochic acid, a commonly ingested herb, and arsenic found in groundwater. Aristolochic death associated protein kinase has also been implicated in endemic Balkan nephropathy and urothelial cancer, and this may have been a risk factor for urothelial cancer in the current series from Basel.
Bacillus Calmette-Guérin (BCG) instillations for the treatment of superficial bladder urothelial cancers in immunocompromised patients are listed on the package insert as a contraindication; however, BCG\’s successful use in transplant recipients has been reported. The use of BCG in this patient population is controversial.

Prostate cancer (PC) is the most common nonskin cancer in men. In 2015, in North America alone, there will be 220,800 new cases of PC and 27,540 men will die of it. PC is also increasing worldwide. There is considerable controversy over prostate-specific antigen-based screening for PC, with no consistent recommendations from major medical organizations on the best approach to screening. While many biomarkers are in development to identify PC, noninvasively using blood and urine assays, the definitive diagnosis of PC relies on the histologic identification of cancer cells on invasive prostate tissue biopsy. The standard biopsy procedure, transrectal ultrasound with 10-12 needle cores of the prostate gland, can be associated with morbidity, and over two-thirds of the time may not identify all malignant lesions.
Radiological examinations such as transrectal ultrasound, X-ray computerized tomography (CT), and multiparametric magnetic resonance imaging, and nuclear scans such as single-photon emission computerized tomography and positron emission tomography (PET), using such radiopharmaceuticals as In-111-ProstaScint, F-18-FDG, and C-11-choline, are available but suffer from limitations. Therefore, there is a compelling need for continuation of the development of a biomolecule, which will detect PC and its metastatic lesions with high sensitivity and specificity.
Recent approaches to imaging PC have generally been directed to targeting prostate-specific membrane antigen (PSMA). We have chosen to target VPAC1, which belongs to the superfamily of G protein-coupled surface receptors that are expressed in high density on certain cancer cells, including PC cells at the onset of oncogenesis and prior to the alterations in cell morphology. VPAC1 receptors (combined for vasoactive intestinal and pituitary adenylate cyclase activating peptide) are involved in cell proliferation, cell differentiations, and survival of PC cells, and are overexpressed in cancer of the prostate, breast, bladder, and lungs. On stroma, normal cells, and benign masses, VPAC1 receptors are minimally present.

These data are consistent with the

These data are consistent with the few published reports of long-term outcomes of RALP. Minnillo et al described a similar success rate of 96.8% in a pediatric chemokine receptor antagonist with a mean age of 10.5 years and mean follow-up of 31.7 months (range 2 to 76 months). In this study, 5 patients required reoperation for persistent UPJ obstruction; 2 secondary to missed crossing vessels, 2 due to recurrent intrinsic stenosis, and 1 for retroperitoneal fibrosis. Mufarrij et al also described a success rate of 95.7% in a cohort of 140 adult patients with a mean follow-up of 29 months (range 3 to 63 months). Six patients required subsequent UPJ procedures, including 2 retrograde laser endopyelotomies of postoperative anastomotic strictures, 2 secondary robotic pyeloplasties, and 2 laparoscopic nephrectomies.
Thus, this study is consistent with others that suggest that robotic assistance in reconstructive procedures such as pyeloplasty produces acceptable results and is the first long-term series to demonstrate that all failures occurred in the first 49 months postoperatively. As minimally invasive pyeloplasty becomes more prevalent and time elapses since the advent of this procedure, future studies may further elucidate risk factors associated with long-term failure. It is suggested in recent literature that, whereas robotic pyeloplasty is currently more expensive than open or purely laparoscopic pyeloplasty due to early investment in robotic equipment costs, RALP may eventually become more cost-effective by reducing inpatient stays and complications once initial investments are made, which may lead to increased utilization in the future. Therefore, it is important for clinicians to be aware of the long-term outcomes of reconstructive procedures such as pyeloplasty when following patients postoperatively.


The authors review their mostly adult patient experience with robot-assisted laparoscopic pyeloplasty (RALP) comprising 129 patients followed for an average of 33.8 months (range 1-147 months). Although their success rate was 96.9%, there were 4 failures, with the failures requiring some treatment after 20 months (2 nephrectomies, 1 long-term stenting, and 1 with long-term suppressive antibiotics). It is noteworthy that 3 of the 4 failures were in patients that did not have a stent for the RALP procedure, although 80.6% were stented at the time of the RALP.

We appreciate the thoughtful comment on the current study. Although perioperative complications are not the emphasis of the current study, it is helpful to analyze them more closely. The urine leak rate was much higher in the stentless group (6/25, 24.0%) when compared to the stented group (3/104, 2.9%,   <  .001). When considering only stented pyeloplasties in our dataset, the 8-year success rate increased to 96.3% from 91.5%. All stentless pyeloplasties were performed prior to 2012, and our current practice is to leave a stent for 4 weeks. Stentless pyeloplasty has been of particular interest in the pediatric population where stent removal might require an anesthetic. In a review of open pediatric stented ( = 339) vs stentless (= 494) pyeloplasty, the overall complication rate was similar (12% vs 14%), respectively. However, secondary procedures were higher in the stentless group (3.5% vs 9.1%,   =  .003). Large or randomized studies on stent use during pyeloplasty have not been performed. Our experience suggests that caution should be used when considering stentless pyeloplasty in the adult population.
Greater than 18 million men in the United States currently suffer from erectile dysfunction (ED), with recent surges in prevalence sustained by increased awareness, availability of effective treatments, and longevity and health in the elderly. Diagnostic and therapeutic strategies in men with ED must balance opportunities to improve sexual quality of life and overall health with the costs of evaluation and treatment. This paradigm has been most extensively studied in cardiovascular disease, which is a common occult cause of morbidity and mortality among men presenting with ED. One recent study suggested that screening men with ED by measurement of blood pressure, serum lipid profile, and hemoglobin A1c could result in an overall healthcare cost savings of >$28 billion over 20 years in the United States alone.

Multiple studies have shown a well established benefit supporting

Multiple studies have shown a well-established benefit supporting lymphadenectomy in patients with urothelial carcinoma undergoing radical cystectomy. The number of TAK-875 nodes removed has been shown to predict cancer-specific mortality, with increasing survival seen with higher lymph node yield. Similarly, improved survival is seen in patients who have undergone an extended lymph node dissection. In addition, lymph node density has been shown to be an independent risk factor for recurrence-free and overall survival. Some studies have even suggested that lymph node density is superior to tumor node metastasis nodal status in predicting cancer-specific survival after radical cystectomy. The survival benefit associated with lymph node yield at the time of radical cystectomy has also been shown to be independent of cancer-positive status of the lymph nodes examined. Thereby, even removing negative lymph nodes provides a survival benefit. One possible explanation for this benefit is that the “negative” lymph nodes that are being removed harbor occult metastatic disease that are being removed, thus, debulking or eliminating the sites of regional disease spread. Our data support this hypothesis, at least in the pT3 population, as we demonstrated occult metastatic urothelial carcinoma in 24% of these patients. Thus, in theory, patients may be benefiting from a thorough debulking of occult metastatic disease.
Incidentally, detected prostate cancer is not an uncommon finding in patients undergoing radical cystoprostatectomy. Studies report an overall incidence of up to 51%, with clinically significant prostate cancer varying from 5% to 60% of these cases. A study by Schiavina et al evaluated the incidence of occult nodal metastases in patients with intermediate and high-risk prostate cancer. Using a combination of serial sectioning, IHC, and real-time reverse transcriptase polymerase chain reaction, they found occult metastatic disease in 33% of patients undergoing prostatectomy, which included 13% additional patients previously considered node negative. In our study, 40% of all male patients had incidentally detected prostate cancer, with 67% of these patients having clinically significant disease. We also discovered occult metastases in 1 patient with pT3a prostate cancer. The discovery of occult metastatic prostate cancer potentially allows patients the option of adjuvant hormonal or radiation therapy; however, the clinical significance of occult metastatic prostate cancer is still unclear, with some studies showing a risk of biochemical recurrence similar to patients with no lymph node metastases.
Extranodal extension and perinodal lymphovascular invasion have been shown to be predictors of clinical outcomes in patients with node-positive bladder cancer treated by cystectomy. In a study of 158 patients with positive lymph nodes, perinodal lymphovascular invasion was TAK-875 an independent prognostic factor for cancer-specific survival. Similarly, in a retrospective analysis of 748 patients with urothelial carcinoma of the bladder treated by radical cystectomy, extranodal extension was associated with disease recurrence and cancer-specific mortality. They also found that the rate of extranodal extension increased with pT stage. In our study, 2 of 5 patients (40%) with occult metastatic urothelial carcinoma had extranodal extension, of which 1 also had perinodal lymphovascular invasion. The patient with both extranodal extension and perinodal lymphovascular invasion had documented metastatic disease at 11 months and died shortly thereafter. This patient also had 3 positive lymph nodes. The second patient with extranodal extension had a solitary micrometastasis and was alive at 29 months without evidence of disease recurrence. Based on our small numbers, it is difficult to comment on the significance of these findings. However, based on other studies, it does represent a prognostic indicator that was missed with routine histologic techniques only to be discovered on deeper levels.

If developing of the talents across our trainees

If developing of the talents across our trainees and young urologists is a goal of urology, then defining what success is for a surgeon, educating on gender bias and its role in divergent female career pathways, and providing active mentorship for female colleagues and young urologists and in their leadership development remain critical issues for this specialty.

The rising prevalence of stone disease in the United States has led to similarly increasing efforts to optimize ureteroscopic treatment. The holmium:yttrium-aluminum-garnet (Ho:YAG) laser, with few exceptions, is the THZ2 source of choice for ureteroscopic therapy because it is safe, efficient, and thoroughly tested. There are several technical factors that influence the performance of holmium laser lithotripsy. Understanding these adjustable elements allows the urologist to enhance their control during laser lithotripsy procedures.
Existing holmium laser lithotripters allow the operator to control the total power output by adjusting pulse energy and pulse frequency. In newer models, the urologist can choose different pulse durations, that is, long pulse vs short pulse. The same amount of energy is delivered per pulse in both settings, but in long pulse mode, the energy is distributed over a longer period of time, reducing retropulsion without sacrificing comminution efficiency.
Previous studies have evaluated the efficiency of variable pulse-width Ho:YAG laser lithotripsy, focusing specifically on the optimal energy and pulse settings. The aim of this study was to perform operator-independent, reproducible experiments to determine differences between the short and long laser pulse duration settings with regard to stone comminution efficiency, laser fiber tip degradation, and retropulsion in a setting designed to mimic the technique of stone dusting.
Materials and Methods


In this study, the long pulse setting on a variable pulse-width laser was shown to offer improved comminution efficiency with less fiber tip degradation and retropulsion than the short pulse setting. Past studies have confirmed that stone fragmentation increases as laser pulse energy increases. However, retropulsion and fiber tip degradation also increase, which combine to decrease overall fragmentation efficiency. By using an adjustable pulse-width laser, the urologist can lengthen the laser pulse duration, thereby overcoming the drawbacks of retropulsion and fiber tip degradation at high power settings, while retaining effective fragmentation capability. This experiment is unique because the comminution model closely mimics the dusting technique that is widely used in clinical practice for stone fragmentation.
Dusting is a fragmentation technique in which the laser tip is brushed back and forth across the stone surface so the THZ2 stone is ablated layer by layer. Uniform erosion of the stone, as opposed to fragmentation, can reduce operative time as fewer large fragments are produced; this decreases the need to search the upper urinary tract for these fragments. The typical dusting method requires a low energy, high frequency laser setting to reduce retropulsion and allow for more uniform distribution of energy across the stone surface. Using long pulse duration allows for a higher energy setting to produce a dusting method of comminution by reducing retropulsion in this way. The model used in our experiment drags a laser fiber tip across the surface of a BegoStone at a constant rate with intermittent adjustment of the fiber tip to keep the fiber tip in close contact with the stone. Unlike other studies, which define comminution efficiency through removal of fragments across a mesh or sieve, our experiment defined comminution efficiency as loss of mass as dust after the stone was treated with a constant amount of laser energy.
Previously, stone comminution using the Swiss LaserClast variable pulse-width laser was evaluated using a standard experimental setup of lattice with a mesh in addition to a hands-free model to exclude experimenter bias. These experimenters found similar comminution efficiency for the short and long pulse widths at a variety of energy settings with less fiber burnback and retropulsion on the long pulse setting. Our study validates these findings using a different experimental model to further demonstrate the benefit of the long pulse, especially when using a dusting technique.

br Materials and methods br Discussion This was a

Materials and methods

This was a hospital-based study investigating the prevalence and predictors of AD in men with LUTS in a medical center in Taiwan. We found that the prevalence of AD was estimated to be 55.7% in men with LUTS in our Urology Outpatient Clinic. There was a considerable portion of patients both with LUTS and AD that had hyperlipidemia, pre-DM, coronary artery disease, and lxr agonists (23.5%, 48.5%, 4.4%, and 46.3%, respectively). Although there was no statistical difference in the prevalence of these associated factors compared with those of the non-AD group, we still need to pay attention to these baseline conditions to identify potential AD patients. As for the other parameters, elevated WBC and WC were significantly associated with AD and increased body weight was borderline associated with AD in the univariate analysis. In the multivariate analysis, however, only men with LUTS with elevated WC revealed a significantly higher risk for AD, and elderly men with LUTS showed a borderline risk for AD. Men with LUTS and WC > 90 cm had a 2.86-fold higher risk for AD than those who had WC < 90 cm. Given that some previous studies found a positive association between metabolic syndrome and LUTS, while other studies reported conflicting data, we only analyzed the parameters in metabolic syndrome (including hyperglycemia, hypertension, triglycerides, high density lipoprotein-C, WC) separately and determined that WC can be seen as an indicator of AD in men with LUTS in Taiwan. Although previous studies have investigated the relationship between AD and LUTS, no consistent correlations were found. In the Third lxr agonists National Health and Nutrition Examination Survey, Rohrmann et al evaluated the association of circulating sex steroid hormone with LUTS. No consistent associations were observed between circulating and free testosterone and risk of LUTS. In Kim et al\’s investigation of the relationship between nocturia and decreased serum testosterone in men with LUTS in Korea, decreased testosterone was a significant independent risk for overall nocturia, especially nocturnal polyuria, and patients with low serum testosterone showed increased nocturnal urine output. Yassin and colleagues proposed the potential role of testosterone in the urinary tract in that testosterone may interact with LUTS via several possible mechanisms, such as by acting on nonpost synaptic receptors in the bladder detrusor muscle and by stimulating nitric oxide production in the urinary tract and bladder. Despite the inconsistencies so far observed in the relationship between AD and LUTS, replacement with testosterone for men with late-onset hypogonadism (LOH) improves LUTS. Testosterone therapy improves LUTS/bladder function by increasing bladder capacity and compliance, and decreasing detrusor pressure in men with LOH. One 5-year prospective, observational study also demonstrated that testosterone replacement is associated with improvements in LUTS and International Prostate Symptom Score in men with LOH. These studies indicate the importance of identifying AD in men with LUTS, with early detection and replacement of testosterone for AD being able to improve the life quality in men with LUTS.
In our study, increased WC rather than body mass index is an independent indicator of AD and in men with LUTS. One epidemiological investigation by Svartberg et al enrolled 1548 men aged 25–84 years surveyed the correlation between sex steroid hormones and central obesity. They found that all hormone associations were stronger for WC than for body mass index and suggested that WC should be the preferred anthropometric measurement in predicting endogenous testosterone levels. Measuring WC is a simple way to predict intra-abdominal fat volume or area and where it is placed around the body and increased intra-abdominal fat is associated with an increase in insulin resistance and systemic inflammation, which may contribute to low testosterone levels. In addition, total and free testosterone levels have been observed to decrease with increasing age in longitudinal studies, and thus it is not surprising in our study that older age was associated with a borderline rise in the risk of AD.

To date no studies have been published examining

To date, no studies have been published examining the shedding kinetics of M genogroup IHNV in rainbow trout over multiple time buy AL 8697 points, or relating this to within host replication kinetics, as done here. Understanding the relationship between replication and shedding is critical because the majority of studies of pathogen buy AL 8697 assume that transmission can be inferred from within host loads, although the association has often not been established (Wargo and Kurath, 2012). In addition, previous studies have generally measured pathogen shedding from fish held in batch, while this study quantified shedding from individual fish, providing insight into both population-based means and fish-to-fish variation in shedding patterns. As such, this study provides a unique evaluation of how viral transmission and fitness vary throughout the infection.



Discussion and conclusions
The replication and shedding kinetics of a pathogen can provide valuable insights into its overall prevalence and transmission in the field, which is vital for informing disease epidemiology and management (LaPatra et al., 2001; Bjornstad et al., 2002; Ogbunugafor et al., 2010; Garver et al., 2013). Studies of these traits can also be used to elucidate how other pathogen traits, such as virulence, might be linked to fitness and thus evolve (Alizon et al., 2009; Wargo and Kurath, 2012). Here, two genotypes of the pathogen IHNV, previously shown to differ in virulence, were examined for their replication and shedding kinetics in rainbow trout. In general, both within host and shed viral loads approached peak values very rapidly, by day 2 post-exposure to virus. Within host cumulative viral loads remained around maximum levels and did not significantly increase or decrease for 3–7days, despite qualitative trends suggesting otherwise (Fig. 1). In contrast, daily viral shedding tapered off quickly, such that it dropped by 2–3 orders of magnitude from day 2 to 5 post-exposure. These kinetics created a peak (day 1–4) and post-peak (days 5 onward) period in shedding, akin to what is seen in other viruses such as HIV (Piatak et al., 1993). This was observed in both the amount of virus shed per fish and the number of fish shedding. The findings here indicate that for IHNV the majority of shedding occurs well before host mortality begins. Whether this is an evolved strategy of the host or the virus in unknown.
The shedding dynamics observed here were different from those found in the only other study of IHNV shedding kinetics, where shedding by infected Atlantic salmon did not become detectable until day 4 and peaked between day 7–10 post-exposure to virus (Garver et al., 2013). There were several differences between our study and this previous one, such as host species, holding temperatures, and use of IHNV strains from different phylogenetic genogroups, all of which are known to influence the replication and virulence of the virus (Garver et al., 2006) and thus potentially shedding. The rapid replication kinetics and early peak viral loads presented here were consistent with a previous study of within host replication of IHNV genotype HV in the same host species O. mykiss (Peñaranda et al., 2009). Collectively, these studies highlight the importance of environmental, host, and virus factors in driving pathogen transmission.
To our knowledge, this is the first published study of IHNV to link replication kinetics in the host with shedding kinetics through time. The data indicated that in general, most of the virus produced is shed into the environment rather than harbored in the host. This suggests that the virus maximizes its fitness by investing heavily in transmission rapidly after infection. However, the ratio of shed to within host virus dropped substantially in the post-peak period of infection, indicating that much less of the virus produced in the host was shed. Why this occurred is unknown. A potential factor could be the destruction of viral RNA within the host by the immune response just prior to packaging and release of virions, perhaps in specific tissues from which shedding occurs. This is supported by the finding that host innate immunity typically peaks and remains at high levels around 48h post-infection (Fig. 2), when shedding begins to decline (Purcell et al., 2010). Shedding may also be limited by host tissue damage, such as necrosis of the kidneys, which inhibits fish urination (Amend and Smith, 1975) and is the main cause of mortality in IHNV infected fish (Bootland and Leong, 2011). This hypothesis is supported by the finding that the post-peak shedding rate reached minimal levels about the time morbidity and mortality began, around day 5 post-infection (Garver et al., 2006). Furthermore, during the post-peak period for shedding, it is unknown if the maintenance of cumulative within host viral load levels at or below maximum levels represents a lack of new viral replication, or if there is continued replication that is counter-balanced by degradation. A decline in active viral RNA replication could contribute to a drop in shedding if only actively replicating or recently synthesized viral genomic RNA is packaged into virions. These potential mechanisms driving the decline in shedding warrant further investigation.

To determine receptor proteins for TMUV co immunoprecipitation assay Co

To determine receptor proteins for TMUV, co-immunoprecipitation assay (Co-IP) was carried out as previously described () with some modifications. The membrane proteins from DF-1 leukotriene receptor antagonist were isolated using the Membrane and Cytosol Protein Extraction Kit (Beyotime, China) (). The DF-1 cell membrane proteins were incubated with purified TMUV on a rocker at 4°C for 6h, followed by 4h leukotriene receptor antagonist of incubation with a mouse specific anti-TMUV E protein polyclonal antibody. After that, protein A-agarose beads were added to the mixture and then incubated for overnight. The beads were washed three times and boiled in 2×SDS loading buffer for 10min. Co-immunoprecipitated complexes were then analyzed by SDS-PAGE gels stained with Coomassie blue. As shown in A, a distinct band with approximate molecular mass 70-kDa was identified in the complexes co-immunoprecipitated with TMUV and anti-TMUV antibody (lane 1), while in the absence of TMUV (lane 3) or anti-TMUV antibody (lane 4), this band was not detected. Thus, we detected a TMUV-binding protein by co-immunoprecipitation assay. To validate this result, virus overlay protein binding assay (VOPBA) was carried out as described earlier (). The co-immunoprecipitated complexes were transferred to nitrocellulose membranes and incubated with TMUV,followed by sequential incubation with a pan specific anti-TMUV E protein monoclonal antibody (), and a goat anti-mouse IgG conjugated with alkaline phosphatase. As shown in B, TMUV recognized the protein of approximate molecular mass 70-kDa (lanes 1), while in the absence of TMUV, the anti-TMUV E protein monoclonal antibody was unable to detect the protein band (lane 2).
To identify the 70-kDa protein, the protein band in gel was excised and sent for commercial mass spectrometry fingerprint. This mass spectrum was compared with protein databases, and the protein was identified as heat shock protein (HSP) A9. Peptide sequences identified by mass spectrometry are shown in C. To further corroborate this identification, the co-immunoprecipitated complexes were transferred to nitrocellulose membranes and probed with a goat anti-HSPA9 antibody (Abcam, UK) which can react with chicken HSPA9, followed by the corresponding secondary antibody conjugated with alkaline phosphatase. As shown in D, the anti-HSPA9 antibody specifically recognized the approximate 70-kDa protein band. Collectively, these results indicated that the HSPA9 in the DF-1 cell membranes could interact with TMUV.
HSPA9, a member of the Hsp70 family of chaperones (also known as Mortalin/GRP75/mtHsp70), is associated multiple cellular functions ranging from stress response, intracellular trafficking, antigen processing, control of cell proliferation, differentiation, and tumorigenesis (). Although originally identified as a mitochondrial chaperone (), HSPA9 is also detected in different subcellular compartments including mitochondria, endoplasmic reticulum, cytoplasmic vesicles, cytosol and plasma membrane (). To determine whether HSPA9 and TMUV colocalized on the cell surface, indirect immunofluorescence assay was performed with non-permeabilized DF-1 cells as described previously (). DF-1 cells in 24-well dishes were infected with TMUV at MOI of 1. After 24h post-infection, the cells were fixed with 3% paraformaldehyde and 2% sucrose in PBS at room temperature for 5min. To check the permeability of the fixed cells, the fixed cells were stained with a mouse anti-β-actin monoclonal antibody (CWBIO, China) as the primary antibody, and a Cy3-labeled goat anti-mouse IgG as the secondary antibody. For analysis of the colocalization of HSPA9 and TMUV, the fixed cells were stained with a goat anti-HSPA9 antibody and a pan specific anti-TMUV E protein monoclonal antibody as described above as the primary antibodies, and an alexa fluor 488-labeled goat anti-rabbit IgG and a Cy3-labeled goat anti-mouse IgG as the secondary antibodies. Nuclei of the cells were stained with DAPI (4, 6-diamidino-2-phenylindole). Visualization under a fluorescent microscope showed a significant cell colocalization between TMUV and HSPA9 (A). Moreover, the anti-β-actin antibody could not detect any molecule on the surface of DF-1 cells under the same conditions (B), which suggests that the integrity of the cells is preserved and TMUV may colocalize with HspA9 on the surface of DF-1 cells.

Fourier transform infrared spectroscopy FTIR is a powerful

Fourier transform infrared spectroscopy (FTIR) is a powerful technique for quantitative and qualitative characterization of biological specimens/molecules in human and veterinary medicine (Shaw and Mantsch, 2000). The use of FTIR for such applications generally flows from the chemometric analysis of the spectroscopic data, which are composed of overlapping phospholipase a2 inhibitor bands that arise from their constituent molecular species (Dubois and Shaw, 2004; Shaw et al., 2008). These absorption patterns for biological tissues and fluids provide the basis to quantify diagnostically relevant constituents in serum, urine and other biological fluid, and may also be used for the direct diagnosis of diseases (Petrich et al., 2002). The possibility of quantitative method development has been realized by the integration of FTIR techniques with chemometric tools such as partial least squares (PLS), principal component analysis, and other approaches. These tools enable the development of quantitative analytical methods that require no molecular separation techniques or reagents (Hou et al., 2014). In our laboratory, these techniques have been previously used as the basis to develop rapid, reagent-free methods for measuring serum or plasma IgG levels in horses and alpacas (Riley et al., 2007; Burns et al., 2014). If the same approach were to provide an accurate test in canines, FTIR spectroscopy may prove to be a desirable testing method for measurement of canine serum (or plasma) IgG concentrations and further investigations into neonatal immunity.

Materials and methods

Of the 193 canines for which a complete signalment was available, 10 were intact females, 76 were spayed females, 24 were intact males, and 83 were neutered males. The mean age of the study population was 6.8±3.9 years (Table 1). The study population encompassed 49 recognized pure breeds of small, medium and large size, as well as a large number of dogs of mixed breeds (Table 1). Of the 193 dogs for which disease status was available, 46 had no clinical illness at the time of sample collection and 147 had disease at sample collection including: 25 with orthopedic disease, 25 with neoplasia, 21 with non-specific gastrointestinal signs, 12 with suspected immune-mediated disease, 11 with renal disease, 10 with diagnosed endocrine disease, 6 with skin allergies, 5 with neurologic signs, 5 with elevated serum biochemistry values consistent with hepatic disease, and 5 with urinary tract infections. The remaining 22 ill animals included individual diseases affecting the integumentary (1), reproductive (2), cardiovascular (4), ophthalmic (1), and respiratory systems (6), as well as urinary sphincter incompetence (2), vehicular trauma (2) and nonspecific clinical signs where no specific diagnosis was obtained (4). The RID IgG concentrations for the 193 serum samples with available signalment data ranged from 432mg/dL to 4979mg/dL. The laboratory reference range for IgG for clinically healthy dogs was 522–4207mg/dL based on the non-parametric option in the Reference Value Advisor (Geffré et al., 2011).
For the FTIR based assay, the 207 samples were sorted based on the IgG concentrations acquired from the RID method. Eight samples with an RID IgG higher than 4000mg/dL were excluded to reduce the leverage effects of this small number of samples, leaving behind spectra from 199 samples for building and validating the analytical method (Fig. 1). The optimal number of PLS factors was determined to be 17 based on the lowest Monte Carlo cross validation value (RMMCCV=529mg/dL). A scatter plot (Fig. 2) exhibits the correlation between IgG concentrations obtained via RID and FTIR (RMSEC=326mg/dL, RMSEP=404mg/dL). The concordance correlation coefficients for the calibration model data set and the prediction set were 0.91 and 0.85, respectively.
The Bland–Altman plot (Fig. 3) shows the mean value of the difference (FTIR–RID) as −89mg/dL. When compared to high IgG concentrations, this value was small and indicated no significant systematic bias between the two methods. A normal probability plot for the differences (Fig. 4) shows the majority of data points scattered closely around the reference line. Precision of the FTIR test and RID analysis is graphically demonstrated in Figs. 5 and 6. The mean coefficient of variation (CV) for RID was 6.67% and for FTIR it was 18.76%.

Introduction Interleukin IL is a member of the

Interleukin-15 (IL-15) is a member of the γ-chain cytokine family, which plays important roles in the regulation of immune responses (Fehniger and Caligiuri, 2001). Binding of IL-15 to its receptor IL-15Rα/IL-2Rβ/γ-chain (γc) generates an immune-enhancing signaling cascade through the JAK/STAT, PI3K/Akt, and Ras/MAPK pathways (Budagian et al., 2006; Lin et al., 1995; Miyazaki et al., 1994; Zambricki et al., 2005; Zhang et al., 2008). These signaling pathways promote the activation, proliferation, and survival of natural killer (NK) Asiatic acid and T cells (Imada et al., 1998; Kelly et al., 2003). More importantly, IL-15 produced by bone marrow and thymus stromal cells stimulates the generation, maintenance, and homeostasis of NK cells (Kennedy et al., 2000). Since IL-15 is essential for generating and activating lymphocyte subsets involved in antitumor immunity, it is considered a promising anticancer immunotherapy drug (Jakobisiak et al., 2011).
IL-15, a critical cytokine for NK cell development, survival and activation, possesses superior advantages for tumor immunotherapy compared to that of IL-2, which also potentiates NK cell function and has been used as an immunotherapeutic agent to promote NK cell antitumor activity (Miller et al., 2014; Pillet et al., 2009). IL-15 has a lower toxicity than IL-2 (Munger et al., 1995), and it supports the survival of NK cells by inhibiting activation-induced cell death (Marks-Konczalik et al., 2000). Unlike IL-2, IL-15 has little effect on the expansion of regulatory T (Treg) cells, which exert an immunosuppressive effect (Berger et al., 2009; Cornish et al., 2006; Zorn et al., 2006). In addition, IL-2, but not IL-15, activated NK cells were shown to increase their sensitivity to apoptosis when these cells come into contact with the vascular endothelium (Rodella et al., 2001). Based on these beneficial features, IL-15 is currently being developed for clinical use as a cancer therapy in humans. This cytokine is also utilized in the ex vivo expansion and activation of NK cells for use in adoptive immunotherapy, and to support the in vivo expansion and function of NK cells after infusion. IL-15 is currently in clinical trials (Miller et al., 2014; Romee et al., 2014).
Only a few attempts have been made to develop a cytokine-based immunotherapy against canine cancer cells, although the incidence of cancers in dogs has increased recently (Chou et al., 2009; Hsiao et al., 2008; Lin et al., 2008). In combination with IL-6 or IL-6 and IL-12, IL-15 has been examined in terms of the antitumor effects exerted through the augmentation of NK cytotoxicity against canine transmissible venereal tumor (CTVT) (Chou et al., 2009; Chuang et al., 2009; Lin et al., 2008). However, most the cytokine genes examined in their reports were from humans. In the present study, we generated an rcIL-15 protein consisting of 114 amino acids (positions 49–162 of the IL-15 preproprotein) for clinical use, and confirmed its biological activity by evaluating its ability to stimulate the proliferation and function of canine non-B, non-T, large granular NK lymphocytes in vitro, which are defined as NK cells in dogs (Knapp et al., 1993). We also determined the ability of rcIL-15 to promote the generation of lymphocytes in vivo after intravenous administration. The results of this study indicated that the purified rcIL-15 could be utilized as a valuable cytokine for cancer immunotherapy, and to support NK cells following adoptive transfer in dogs.

Materials and methods


IL-15 is a promising cytokine with the greatest potential for use in tumor immunotherapy because it plays crucial roles in antitumor mechanisms by promoting the generation, activation, proliferation, and survival of lymphocyte subsets, including NK cells, CD8+ T cells, and NKT cells (Jakobisiak et al., 2011). IL-15, which is known as an essential mediator of NK cell homeostasis, is effective for treating a variety of Asiatic acid tumors in animal models (Steel et al., 2012), and its safety has been evaluated in mice (Munger et al., 1995) and in non-human primates (Mueller et al., 2005). In addition, clinical trials using rhIL-15 alone or in combination with other drugs to treat various types of human cancer have been carried out (Steel et al., 2012; Wu, 2013). Moreover, IL-15 has been extensively exploited as a powerful anticancer drug; e.g., as a superagonist combined with modified IL-15Rα. In contrast, no attempts have been made to use the canine IL-15 protein to treat cancer in dogs. Due to the important roles of IL-15 in antitumor immunity, the rcIL-15 protein may be useful in canine tumor immunotherapy.

RepSox Increased prevalence of bacterial strains resistant to

Increased prevalence of bacterial strains resistant to RepSox in humans has stimulated public and federal interest in eliminating the use of antibiotics in sub-therapeutic doses for growth promotion (antibiotic-growth promoters; AGP) in livestock. An alternative approach to improve health and productivity in livestock is the use of probiotics, prebiotic substrates that serve as nutrients to certain bacteria, or their combinations (synbiotics). A variety of microbial species (bacteria of Bacillus, Escherichia, Lactobacillus, Bifidobacterium, Enterococcus, Lactococcus, Streptococcus, and Pediococcus genera, yeast and undefined mixed cultures) have been used as probiotics generally resulting in reduced mortality, enhanced immune responses, improved growth rates, feed intake and feed efficiency in poultry and livestock of different ages [reviewed in Cho et al. (2011) and Patterson and Burkholder (2003)]. While Lactobacillus and Bifidobacterium species have been used most extensively in humans; historically, various species of Bacillus, Enterococcus, and Saccharomyces yeast have been the most commonly used in livestock (Simon et al., 2001). Only during the past few decades, has there been an increase in research on supplementing Lactobacillus to livestock (Gusils et al., 1999; Jin et al., 2000; Pascual et al., 1999; Tellez et al., 2001) (Table 3). Further, while in some studies LAPB improved growth performance and post-weaning diarrhea (PWD) control in weanling pigs (Lessard and Brisson, 1987; Shu et al., 2001), these effects were not observed in others (Walsh et al., 2007) (Table 3). As reviewed in Heo et al. (2013), this inconsistency in results of probiotic effects on PWD and performance in pigs may be attributed to differences in dosage and type of probiotic, management practices, diet, and age (Heo et al., 2013). One study evaluated the effects of bifidobacteria and LAPB (in place of AGPs) in newborn calves and piglets and demonstrated that these probiotics reduced mortality, improved weight gain, fecal condition and feed efficiency in both species (Abe et al., 1995). However, the effects of lactobacilli (including various strains of L. reuteri, as well as Lactobacillus gasseri, L. acidophilus and L. fermentum) supplementation on infectious diarrhea occurrence, growth performance and feed conversion in neonatal and weanling piglets varied with age, feeding status (sow milk versus milk replacer) and lactobacilli strain (Chang et al., 2001; Chen et al., 2014; Huang et al., 2004; Liu et al., 2014; Wang et al., 2009a, 2013, 2011, 2012; Yu et al., 2008) (Table 3). Potential mechanisms of lactobacilli beneficial effects proposed in these studies included alleviation of oxidative stress (Wang et al., 2013, 2009b), protective modulation of gut microbiota (Chang et al., 2001; Huang et al., 2004; Liu et al., 2014) and associated metabolic profiles (Liu et al., 2014), enhancement of T-cell differentiation, ileal cytokine production (Wang et al., 2009a) and serum IgG Ab levels (Yu et al., 2008). Additionally, reduction in the levels of IL-1β mRNA expression in the ileum of neonatal piglets due to L. reuteri supplementation was reported (Hou et al., 2015; Liu et al., 2014).
Very few mechanistic studies addressing interactions among LAPB, immunity and RV were conducted in livestock species, and primarily in pigs. In 3-week old piglets, the administration of B. lactis HN019 led to lower concentrations of fecal RVA and reduced severity of weanling diarrhea (Shu et al., 2001) (Table 3). Indicative of immune enhancement, higher blood leukocyte phagocytic and T-lymphocyte proliferative responses, and higher intestinal RV-specific Ab (IgM, IgG and IgA) titers were detected in B. lactis HN019 fed piglets. Interestingly, another study using suckling piglets demonstrated reduced RVA shedding due to Enterococcus faecium NCIMB 10415 supplementation that was not associated with increased RV-specific Ab titers (Kreuzer et al., 2012). However, the probiotic supplementation resulted in significant differences in effector and regulatory T cell responses. These data suggest, once again that reduction in RV diarrhea/infection may be achieved via different mechanisms by different probiotic bacteria, while the increase of RVA-specific Ab levels (often found due to probiotic supplementation) is not essential for the disease attenuation.