Programmed death 1 signaling on chronic myeloid leukemia-specific T cells results in T-cell exhaustion and disease progression

Chronic myeloid leukemia (CML) is a malignant myeloproliferative disease with a characteristic chronic phase (cp) of several years before progression to blast crisis (bc). The immune system may contribute to disease control in CML. We analyzed leukemia-specific immune responses in cpCML and bcCML in a retroviral-induced murine CML model. In the presence of cpCML and bcCML expressing the glycoprotein of lymphocytic choriomeningitis virus as a model leukemia antigen, leukemia-specific cytotoxic T lymphocytes (CTLs) became exhausted. They maintained only limited cytotoxic activity, and did not produce interferon-gamma or tumor necrosis factor-alpha or expand after restimulation. CML-specific CTLs were characterized by high expression of programmed death 1 (PD-1), whereas CML cells expressed PD-ligand 1 (PD-L1). Blocking the PD-1/PD-L1 interaction by generating bcCML in PD-1-deficient mice or by repetitive administration of alphaPD-L1 antibody prolonged survival. In addition, we found that PD-1 is up-regulated on CD8(+) T cells from CML patients. Taken together, our results suggest that blocking the PD-1/PD-L1 interaction may restore the function of CML-specific CTLs and may represent a novel therapeutic approach for CML.

Perturbation of phosphatidylethanolamine synthesis affects mitochondrial morphology and cell-cycle progression in procyclic-form Trypanosoma brucei

Phosphatidylethanolamine (PE) and phosphatidylcholine (PC) are the two major constituents of eukaryotic cell membranes. In the protist Trypanosoma brucei, PE and PC are synthesized exclusively via the Kennedy pathway. To determine which organelles or processes are most sensitive to a disruption of normal phospholipid levels, the cellular consequences of a decrease in the levels of PE or PC, respectively, were studied following RNAi knock-down of four enzymes of the Kennedy pathway. RNAi against ethanolamine-phosphate cytidylyltransferase (ET) disrupted mitochondrial morphology and ultrastructure. Electron microscopy revealed alterations of inner mitochondrial membrane morphology, defined by a loss of disk-like cristae. Despite the structural changes in the mitochondrion, the cells maintained oxidative phosphorylation. Our results indicate that the inner membrane morphology of T. brucei procyclic forms is highly sensitive to a decrease of PE levels, as a change in the ultrastructure of the mitochondrion is the earliest phenotype observed after RNAi knock-down of ET. Interference with phospholipid synthesis also impaired normal cell-cycle progression. ET RNAi led to an accumulation of multinucleate cells. In contrast, RNAi against choline-/ethanolamine phosphotransferase, which affected PC as well as PE levels, caused a cell division phenotype characterized by non-division of the nucleus and production of zoids.

Region Specific Response of Intervertebral Disc Cells to Complex Dynamic Loading: An Organ Culture Study Using a Dynamic Torsion-Compression Bioreactor

The spine is routinely subjected to repetitive complex loading consisting of axial compression, torsion, flexion and extension. Mechanical loading is one of the important causes of spinal diseases, including disc herniation and disc degeneration. It is known that static and dynamic compression can lead to progressive disc degeneration, but little is known about the mechanobiology of the disc subjected to combined dynamic compression and torsion. Therefore, the purpose of this study was to compare the mechanobiology of the intervertebral disc when subjected to combined dynamic compression and axial torsion or pure dynamic compression or axial torsion using organ culture. We applied four different loading modalities 1. control: no loading (NL), 2. cyclic compression (CC), 3. cyclic torsion (CT), and 4. combined cyclic compression and torsion (CCT) on bovine caudal disc explants using our custom made dynamic loading bioreactor for disc organ culture. Loads were applied for 8 h/day and continued for 14 days, all at a physiological magnitude and frequency. Our results provided strong evidence that complex loading induced a stronger degree of disc degeneration compared to one degree of freedom loading. In the CCT group, less than 10\% nucleus pulposus (NP) cells survived the 14 days of loading, while cell viabilities were maintained above 70\% in the NP of all the other three groups and in the annulus fibrosus (AF) of all the groups. Gene expression analysis revealed a strong up-regulation in matrix genes and matrix remodeling genes in the AF of the CCT group. Cell apoptotic activity and glycosaminoglycan content were also quantified but there were no statistically significant differences found. Cell morphology in the NP of the CCT was changed, as shown by histological evaluation. Our results stress the importance of complex loading on the initiation and progression of disc degeneration.

Differences in Time to Disease Progression Do Not Predict for Cancer-specific Survival in Patients Receiving Immediate or Deferred Androgen-deprivation Therapy for Prostate Cancer: Final Results of EORTC Randomized Trial 30891 with 12 Years of Follow-up

BACKGROUND Trials assessing the benefit of immediate androgen-deprivation therapy (ADT) for treating prostate cancer (PCa) have often done so based on differences in detectable prostate-specific antigen (PSA) relapse or metastatic disease rates at a specific time after randomization. OBJECTIVE Based on the long-term results of European Organization for Research and Treatment of Cancer (EORTC) trial 30891, we questioned if differences in time to progression predict for survival differences. DESIGN, SETTING, AND PARTICIPANTS EORTC trial 30891 compared immediate ADT (n=492) with orchiectomy or luteinizing hormone-releasing hormone analog with deferred ADT (n=493) initiated upon symptomatic disease progression or life-threatening complications in randomly assigned T0-4 N0-2 M0 PCa patients. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS Time to first objective progression (documented metastases, ureteric obstruction, not PSA rise) and time to objective castration-resistant progressive disease were compared as well as PCa mortality and overall survival. RESULTS AND LIMITATIONS After a median of 12.8 yr, 769 of the 985 patients had died (78%), 269 of PCa (27%). For patients receiving deferred ADT, the overall treatment time was 31% of that for patients on immediate ADT. Deferred ADT was significantly worse than immediate ADT for time to first objective disease progression (p<0.0001; 10-yr progression rates 42% vs 30%). However, time to objective castration-resistant disease after deferred ADT did not differ significantly (p=0.42) from that after immediate ADT. In addition, PCa mortality did not differ significantly, except in patients with aggressive PCa resulting in death within 3-5 yr after diagnosis. Deferred ADT was inferior to immediate ADT in terms of overall survival (hazard ratio: 1.21; 95% confidence interval, 1.05-1.39; p [noninferiority]=0.72, p [difference] = 0.0085). CONCLUSIONS This study shows that if hormonal manipulation is used at different times during the disease course, differences in time to first disease progression cannot predict differences in disease-specific survival. A deferred ADT policy may substantially reduce the time on treatment, but it is not suitable for patients with rapidly progressing disease.

Factors influencing the onset and progression of pododermatitis in captive flamingos (Phoenicopteridae).

Pododermatitis is a worldwide problem in captive flamingos. We performed an evaluation of different influence factors (age, sex, weight, origin, breeding status) and a comparison of foot lesions between several zoological institutions and the feet of free-ranging Greater flamingos (Phoenicopterus roseus). A scoring system was used to determine the prevalence and types of lesions and severity. Cracks and nodules developed as early as 3 months of age and papillomatous growths as early as 6 to 7 months of age in captivity. Nodules with ulceration occurred significantly more often in birds older than 31 years and heavier than 4 kg. The comparison of different institutions revealed that birds kept in enclosures with natural-floored water ponds had significantly less severe lesions than birds kept in concrete water ponds. None of the free-ranging flamingos, which live on a muddy underground, showed any lesion. This study demonstrates that flooring, weight and age are important in the onset and progression of pododermatitis in flamingos.

Intraductal papillary neoplasms of the bile duct: stepwise progression to carcinoma involves common molecular pathways

Intraductal papillary neoplasms of the bile duct are still poorly characterized regarding (1) their molecular alterations during the development to invasive carcinomas, (2) their subtype stratification and (3) their biological behavior. We performed a multicenter study that analyzed these issues in a large European cohort. Intraductal papillary neoplasms of the bile duct from 45 patients were graded and subtyped using mucin markers and CDX2. In addition, tumors were analyzed for common oncogenic pathways, and the findings were correlated with subtype and grade. Data were compared with those from 22 extra- and intrahepatic cholangiocarcinomas. Intraductal papillary neoplasms showed a development from preinvasive low- to high-grade intraepithelial neoplasia to invasive carcinoma. Molecular and immunohistochemical analysis revealed mutated KRAS, overexpression of TP53 and loss of p16 in low-grade intraepithelial neoplasia, whereas loss of SMAD4 was found in late phases of tumor development. Alterations of HER2, EGFR, β-catenin and GNAS were rare events. Among the subtypes, pancreato-biliary (36%) and intestinal (29%) were the most common, followed by gastric (18%) and oncocytic (13%) subtypes. Patients with intraductal papillary neoplasm of the bile duct showed a slightly better overall survival than patients with cholangiocarcinoma (hazard ratio (cholangiocarcinoma versus intraductal papillary neoplasm of the bile duct): 1.40; 95% confidence interval: 0.46-4.30; P=0.552). The development of biliary intraductal papillary neoplasms of the bile duct follows an adenoma-carcinoma sequence that correlates with the stepwise activation of common oncogenic pathways. Further large trials are needed to investigate and verify the finding of a better prognosis of intraductal papillary neoplasms compared with conventional cholangiocarcinoma.

Oxygen regulates molecular mechanisms of cancer progression and metastasis

Oxygen is the basic molecule which supports life and it truly is “god's gift to life.” Despite its immense importance, research on “oxygen biology” has never received the light of the day and has been limited to physiological and biochemical studies. It seems that in modern day biology, oxygen research is summarized in one word “hypoxia.” Scientists have focused on hypoxia-induced transcriptomics and molecular–cellular alterations exclusively in disease models. Interestingly, the potential of oxygen to control the basic principles of biology like homeostatic maintenance, transcription, replication, and protein folding among many others, at the molecular level, has been completely ignored. Here, we present a perspective on the crucial role played by oxygen in regulation of basic biological phenomena. Our conclusion highlights the importance of establishing novel research areas like oxygen biology, as there is great potential in this field for basic science discoveries and clinical benefits to the society.

Hierarchical accumulation of RyR post-translational modifications drives disease progression in dystrophic cardiomyopathy

AIMS:Duchenne muscular dystrophy (DMD) is a muscle disease with serious cardiac complications. Changes in Ca(2+) homeostasis and oxidative stress were recently associated with cardiac deterioration, but the cellular pathophysiological mechanisms remain elusive. We investigated whether the activity of ryanodine receptor (RyR) Ca(2+) release channels is affected, whether changes in function are cause or consequence and which post-translational modifications drive disease progression. METHODS AND RESULTS:Electrophysiological, imaging, and biochemical techniques were used to study RyRs in cardiomyocytes from mdx mice, an animal model of DMD. Young mdx mice show no changes in cardiac performance, but do so after ∼8 months. Nevertheless, myocytes from mdx pups exhibited exaggerated Ca(2+) responses to mechanical stress and 'hypersensitive' excitation-contraction coupling, hallmarks of increased RyR Ca(2+) sensitivity. Both were normalized by antioxidants, inhibitors of NAD(P)H oxidase and CaMKII, but not by NO synthases and PKA antagonists. Sarcoplasmic reticulum Ca(2+) load and leak were unchanged in young mdx mice. However, by the age of 4-5 months and in senescence, leak was increased and load was reduced, indicating disease progression. By this age, all pharmacological interventions listed above normalized Ca(2+) signals and corrected changes in ECC, Ca(2+) load, and leak. CONCLUSION:Our findings suggest that increased RyR Ca(2+) sensitivity precedes and presumably drives the progression of dystrophic cardiomyopathy, with oxidative stress initiating its development. RyR oxidation followed by phosphorylation, first by CaMKII and later by PKA, synergistically contributes to cardiac deterioration.

Multidrug resistance protein 4 (MRP4) expression in prostate cancer is associated with androgen signaling and decreases with tumor progression

Multidrug resistance protein 4 (MRP4) is a transmembrane transport protein found in many cell types and is involved in substrate-specific transport of endogenous and exogenous substrates. Recently, it has shown to be expressed in prostate cancer cell lines and to be among the most commonly upregulated transcripts in prostate cancer, although a comprehensive expression analysis is lacking so far. We aimed to investigate its expression by immunohistochemistry in a larger cohort of neoplastic and nonneoplastic prostate tissues (n = 441) and to correlate its expression with clinicopathological parameters including PSA-free survival times and molecular correlates of androgen signaling (androgen receptor (AR), prostate-specific antigen (PSA), and forkhead box A (FoxA)). MRP4 is widely expressed in benign and neoplastic prostate epithelia, but its expression gradually decreases during tumor progression towards castrate-resistant disease. Concordantly, it correlated with conventional prognosticators of disease progression and-within the group of androgen-dependent tumors-with AR and FoxA expression. Moreover, lower levels of MRP4 expression were associated with shorter PSA relapse-free survival times in the androgen-dependent group. In benign tissues, we found zone-dependent differences of MRP4 expression, with the highest levels in the peripheral and central zones. Although MRP4 is known to be regulated in prostate cancer, this study is the first to demonstrate a gradual downregulation of MRP4 protein during malignant tumor progression and a prognostic value of this loss of expression.

Highly pathogenic adapted HIV-1 strains limit host immunity and dictate rapid disease progression.

OBJECTIVE: The study of HIV-1 rapid progressors has been limited to specific case reports. Nevertheless, identification and characterization of the viral and host factors involved in rapid progression are crucial when attempting to uncover the correlates of rapid disease outcome. DESIGN: We carried out comparative functional analyses in rapid progressors (n = 46) and standard progressors (n = 46) early after HIV-1 seroconversion (≤1 year). The viral traits tested were viral replicative capacity, co-receptor usage, and genomic variation. Host CD8 T-cell responses, humoral activity, and HLA immunogenetic markers were also determined. RESULTS: Our data demonstrate an unusual convergence of highly pathogenic HIV-1 strains in rapid progressors. Compared with standard progressors, rapid progressor viral strains show higher in-vitro replicative capacity (81.5 vs. 67.9%; P = 0.025) and greater X4/DM co-receptor usage (26.3 vs. 2.8%; P = 0.006) in early infection. Limited or absent functional HIV-1 CD8 T-cell responses and neutralizing activity were measured in rapid progressors. Moreover, the increase in common HLA allele-restricted CD8 T-cell escape mutations in rapid progressors acts as a signature of uncontrolled HIV-1 replication and early impairment of adaptive cellular responses. CONCLUSION: Our data support a dominant role for viral factors in rapid progressors. Robust HIV-1 replication and intrinsic viral properties limit host adaptive immune responses, thus driving rapid disease progression.

The craniosacral progression of muscle development influences the emergence of neuromuscular junction alterations in a severe murine model for spinal muscular atrophy.

AIMS As 4-day-old mice of the severe spinal muscular atrophy (SMA) model (dying at 5-8 days) display pronounced neuromuscular changes in the diaphragm but not the soleus muscle, we wanted to gain more insight into the relationship between muscle development and the emergence of pathological changes and additionally to analyse intercostal muscles which are affected in human SMA. METHODS Structures of muscle fibres and neuromuscular junctions (NMJs) of the diaphragm, intercostal and calf muscles of prenatal (E21) and postnatal (P0 and P4) healthy and SMA mice were analysed by light and transmission electron microscopy. NMJ innervation was studied by whole mount immunofluorescence in diaphragms of P4 mice. RESULTS During this period, the investigated muscles still show a significant neck-to-tail developmental gradient. The diaphragm and calf muscles are most and least advanced, respectively, with respect to muscle fibre fusion and differentiation. The number and depth of subsynaptic folds increases, and perisynaptic Schwann cells (PSCs) acquire a basal lamina on their outer surface. Subsynaptic folds are connected to an extensive network of tubules and beaded caveolae, reminiscent of the T system in adult muscle. Interestingly, intercostal muscles from P4 SMA mice show weaker pathological involvement (that is, vacuolization of PSCs and perineurial cells) than those previously described by us for the diaphragm, whereas calf muscles show no pathological changes. CONCLUSION SMA-related alterations appear to occur only when the muscles have reached a certain developmental maturity. Moreover, glial cells, in particular PSCs, play an important role in SMA pathogenesis.

Disease progression and recurrence in women treated for vulvovaginal intraepithelial neoplasia

OBJECTIVE The malignant potential of intraepithelial neoplasia of the vulva and vagina after treatment is not well defined. Our objective was to examine risk factors for recurrence and invasive disease. METHODS Four hundred sixty-four women with biopsy proven high-grade intraepithelial neoplasia of the vulva and vagina were identified in the electronic databases of four colposcopy clinics. Inclusion criteria were a follow-up of more than one year, no history of invasive cancer and no invasive cancer within the first year after initial treatment. We investigated the potential factors associated with recurrence and progression using a logistic regression analysis to estimate odds ratios (ORs) and 95% confidence intervals (CIs). RESULTS Of the 411 eligible patients, 123 patients (29.9%) recurred later than one year after initial treatment and 24 patients (5.8%) progressed to invasive disease. According to multivariate analyses, the risk factors associated with recurrence were multifocality (OR, 3.33; 95% CI, 2.02 to 5.51), immunosuppression (OR, 2.51; 95% CI, 1.09 to 5.81), excision as initial treatment (vs. laser evaporation; OR, 1.79; 95% CI, 1.11 to 2.91) and smoking (OR, 1.61; 95% CI, 1.02 to 2.55). Risk factors for progression to invasive disease were immunosuppression (OR, 4.00; 95% CI, 1.30 to 12.25), multifocality (OR, 3.05; 95% CI, 1.25 to 7.43) and smoking (OR, 2.97; 95% CI, 1.16 to 7.60), but not treatment modality. CONCLUSION Laser evaporation combined with extensive biopsy is at least as efficacious as initial treatment of intraepithelial neoplasia with excision. Smoking is a risk factor for both recurrence and progression to invasive disease. Hence, smoking cessation should be advised and maintaining a long follow-up period due to late relapses is necessary.

Latrepirdine improves cognition and arrests progression of neuropathology in an Alzheimer's mouse model

Latrepirdine (Dimebon) is a pro-neurogenic, antihistaminic compound that has yielded mixed results in clinical trials of mild to moderate Alzheimer's disease, with a dramatically positive outcome in a Russian clinical trial that was unconfirmed in a replication trial in the United States. We sought to determine whether latrepirdine (LAT)-stimulated amyloid precursor protein (APP) catabolism is at least partially attributable to regulation of macroautophagy, a highly conserved protein catabolism pathway that is known to be impaired in brains of patients with Alzheimer's disease (AD). We utilized several mammalian cellular models to determine whether LAT regulates mammalian target of rapamycin (mTOR) and Atg5-dependent autophagy. Male TgCRND8 mice were chronically administered LAT prior to behavior analysis in the cued and contextual fear conditioning paradigm, as well as immunohistological and biochemical analysis of AD-related neuropathology. Treatment of cultured mammalian cells with LAT led to enhanced mTOR- and Atg5-dependent autophagy. Latrepirdine treatment of TgCRND8 transgenic mice was associated with improved learning behavior and with a reduction in accumulation of Aβ42 and α-synuclein. We conclude that LAT possesses pro-autophagic properties in addition to the previously reported pro-neurogenic properties, both of which are potentially relevant to the treatment and/or prevention of neurodegenerative diseases. We suggest that elucidation of the molecular mechanism(s) underlying LAT effects on neurogenesis, autophagy and behavior might warranty the further study of LAT as a potentially viable lead compound that might yield more consistent clinical benefit following the optimization of its pro-neurogenic, pro-autophagic and/or pro-cognitive activities.