The role of pro-domains in human growth factors and cytokines.

Many growth factors and cytokines are produced as larger precursors, containing pro-domains, that require proteolytic processing to release the bioactive ligand. These pro-domains can be significantly larger than the mature domains and can play an active role in the regulation of the ligands. Mining the UniProt database, we identified almost one hundred human growth factors and cytokines with pro-domains. These are spread across several unrelated protein families and vary in both their size and composition. The precise role of each pro-domain varies significantly between the protein families. Typically they are critical for controlling bioactivity and protein localisation, and they facilitate diverse mechanisms of activation. Significant gaps in our understanding remain for pro-domain function - particularly their fate once the bioactive ligand has been released. Here we provide an overview of pro-domain roles in human growth factors and cytokines, their processing, regulation and activation, localisation as well as therapeutic potential.

A synthetic synaptic organizer protein restores glutamatergic neuronal circuits.

Neuronal synapses undergo structural and functional changes throughout life, which are essential for nervous system physiology. However, these changes may also perturb the excitatory-inhibitory neurotransmission balance and trigger neuropsychiatric and neurological disorders. Molecular tools to restore this balance are highly desirable. Here, we designed and characterized CPTX, a synthetic synaptic organizer combining structural elements from cerebellin-1 and neuronal pentraxin-1. CPTX can interact with presynaptic neurexins and postsynaptic AMPA-type ionotropic glutamate receptors and induced the formation of excitatory synapses both in vitro and in vivo. CPTX restored synaptic functions, motor coordination, spatial and contextual memories, and locomotion in mouse models for cerebellar ataxia, Alzheimer's disease, and spinal cord injury, respectively. Thus, CPTX represents a prototype for structure-guided biologics that can efficiently repair or remodel neuronal circuits.

Transplantation of Mesenchymal Stromal Cells Expressing the Human Preproenkephalin Gene Can Relieve Pain in a Rat Model of Neuropathic Pain.

Transgenic therapy for central neuralgia faces the problems of low expression and weak targeting and affects superficial but not deep neurons. In this study, we generated a lentivirus vector with human preproenkephalin gene (hPPE) expression driven by the transcriptional amplification strategy system (TAS) and established a primary bone marrow-derived mesenchymal stromal cell (BMSC) line stably expressing hPPE for transplantation into a rat model of neuropathic pain rat. The paw thermal withdrawal latency assay and paw mechanical withdrawal threshold assay showed that unlike control BMSCs and BMSCs with hPPE overexpression driven by the CMV or Synapsin 1 (SYN1) promoter, TAS-hPPE BMSCs had a robust and lasting analgesic effect. The TAS-hPPE BMSC-treated group exhibited higher expression of TAS-driven hPPE and a higher ratio of BMSCs in the midbrain, spinal cord and cortex then the CMV-hPPE BMSC- and SYN1-hPPE BMSC-treated groups. Moreover, we also observed that TAS-hPPE BMSCs displayed a greater tendency to differentiate into neurons and exhibit neuronal-like distribution than CMV-hPPE or SYN1-hPPE BMSCs. In conclusion, our study shows that the TAS improves BMSC transgenic therapy for neuropathic pain treatment.

Gestational diabetes insipidus: Diagnosis and management.

In the pregnant patient, hypotonic polyuria in the setting of elevated serum osmolality and polydipsia should narrow the differential to causes related to diabetes insipidus (DI). Gestational DI, also called transient DI of pregnancy, is a distinct entity, unique from central DI or nephrogenic DI which may both become exacerbated during pregnancy. These three different processes relate to vasopressin, where increased metabolism, decreased production or altered renal sensitivity to this neuropeptide should be considered. Gestational DI involves progressively rising levels of placental vasopressinase throughout pregnancy, resulting in decreased endogenous vasopressin and resulting hypotonic polyuria worsening through the pregnancy. Gestational DI should be distinguished from central and nephrogenic DI that may be seen during pregnancy through use of clinical history, urine and serum osmolality measurements, response to desmopressin and potentially, the newer, emerging copeptin measurement. This review focuses on a brief overview of osmoregulatory and vasopressin physiology in pregnancy and how this relates to the clinical presentation, pathophysiology, diagnosis and management of gestational DI, with comparisons to the other forms of DI during pregnancy. Differentiating the subtypes of DI during pregnancy is critical in order to provide optimal management of DI in pregnancy and avoid dehydration and hypernatremia in this vulnerable population.

Dual-targeting nanoparticle vaccine elicits a therapeutic antibody response against chronic hepatitis B.

Chronic hepatitis B is caused by prolonged infection with the hepatitis B virus (HBV), which can substantially increase the risk of developing liver disease. Despite the development of preventive vaccines against HBV, a therapeutic vaccine inducing an effective antibody response still remains elusive. The preS1 domain of the large HBV surface protein is the major viral attachment site on hepatocytes and thus offers a therapeutic target; however, its poor immunogenicity limits clinical translation. Here, we design a ferritin nanoparticle vaccine that can deliver preS1 to specific myeloid cells, including SIGNR1+ dendritic cells (which activate T follicular helper cells) and lymphatic sinus-associated SIGNR1+ macrophages (which can activate B cells). This nanoparticle vaccine induces a high-level and persistent anti-preS1 response that results in efficient viral clearance and partial serological conversion in a chronic HBV mouse model, offering a promising translatable vaccination strategy for the functional cure of chronic hepatitis B.

Prothymosin alpha and its mimetic hexapeptide improve delayed tissue plasminogen activator-induced brain damage following cerebral ischemia.

Tissue plasminogen activator (tPA) administration beyond 4.5 h of stroke symptoms is beneficial for patients but has an increased risk of cerebral hemorrhage. Thus, increasing the therapeutic window of tPA is important for stroke recovery. We previously showed that prothymosin alpha (ProTα) or its mimetic hexapeptide (P6Q) has anti-ischemic activity. Here, we examined the beneficial effects of ProTα or P6Q against delayed tPA-induced brain damage following middle cerebral artery occlusion (MCAO) or photochemically induced thrombosis in mice. Brain hemorrhage was observed by tPA administration during reperfusion at 4.5 and 6 h after MCAO. Co-administration of ProTα with tPA at 4.5 h inhibited hemorrhage and motor dysfunction 2-4 days, but not 7 days after MCAO. ProTα administration at 2 and 4.5 h after MCAO significantly inhibited tPA (4.5 h)-induced motor dysfunction and death more than 7 days. Administration of tPA caused the loss of tight junction proteins, zona occulden-1 and occludin, and up-regulation of matrix metalloproteinase-2/9, in a ProTα-reversible manner. P6Q administration abolished tPA (4.5 h)-induced hemorrhage and reversed tPA (6 h)-induced vascular damage and matrix metalloproteinase-2 and 9 up-regulation. Twice administrations of P6Q at 2 h alone and 6 h with tPA significantly improved motor dysfunction more than 7 days. In photochemically induced thrombosis ischemia, similar vascular leakage and neuronal damage (infarction and motor dysfunction) by late tPA (4.5 or 6 h) were also inhibited by P6Q. Thus, these studies suggest that co-administration with ProTα or P6Q would be beneficial to inhibit delayed tPA-induced hemorrhagic mechanisms in acute ischemic stroke.

Procalcitonin and MR-proAdrenomedullin combination in the etiological diagnosis and prognosis of sepsis and septic shock.

Procalcitonin and Mid-regional pro Adrenomedullin have been proposed for sepsis diagnosis, antibiotic therapy guidance and prognosis. A retrospective analysis of PCT and MR-proADM on 571 consecutive patients with sepsis diagnosis was performed. Median values were compared using the non-parametric Mann-Whitney's test. Receiver operating characteristic analysis was performed to define cutoff points for sepsis diagnosis. Pretest odds, posttest odds, and posttest probability have been calculated. Data were analyzed using Med-Calc 11.6.1.0 software. PCT resulted excellent in gram-negative, but less performant in gram-positive and fungal etiologies. MR-proADM values resulted homogenously distributed within the different microbial classes and increased significantly in septic shock. PCT highest PPV value was found to distinguish gram-negative from fungal sepsis and septic shock (>3. 57 ng/mL, PPV 0.96 and > 8.77 ng/mL, PPV 0.96, respectively). Good diagnostic accuracy was evidenced to discriminate gram-negative from gram-positive septic shock (>3.88 ng/mL PPV 0.89). Lower diagnostic accuracy was evidenced to discriminate gram-negative and gram-positive sepsis (>0.80 ng/mL, PPV 0.78) and gram-positive from fungal septic shock (>1.74 ng/mL PPV 0.75). The lowest PCT PPV (0.28) was found in gram-positive and fungal sepsis distinction. MR-proADM discriminating cut-offs were homogeneously distributed in Gram-negative and Gram-positive sepsis and were higher in septic shock, but not influenced by pathogen etiologies. MR-proADM cut-off values > 3.39 nmol/L in sepsis and >4.33 nmol/L in septic shock were associated with significant higher risk of 90-days mortality. In conclusion, PCT and MR-proADM combination represents an advantage for sepsis diagnosis and for 90-days mortality risk stratification.

proBDNF Accelerates Brain Amyloid-ß Deposition and Learning and Memory Impairment in APPswePS1dE9 Transgenic Mice.

BACKGROUND: Alzheimer's disease (AD) is pathologically known for the amyloid-ß (Aß) deposition, neurofibrillary tangles, and neuronal loss in the brain. The precursor of brain-derived neurotrophic factor (proBDNF) before proteolysis has opposing functions to its mature form in neuronal survival and neurite growth. However, the role of proBDNF in the pathogenesis of AD remains unclear. OBJECTIVE: To investigate the effects of proBDNF on neurons in vitro, and on learning and memory impairment and brain Aß production in a transgenic AD mouse model (APPswePS1dE9). METHODS: We here examined the effects of proBDNF on the viability (MTT assay) and neurite growth (morphologic measurement) of the primary neurons in vitro. After the intracerebroventricular injection of adeno-associated virus-proBDNF (AAV-proBDNF), we then investigated the learning and memory impairment (Morris water maze) and Aß deposition in the brains of the AD mice. RESULTS: The results showed that proBDNF could inhibit neuronal viability and neurite growth in vitro, enhance Aß levels, and accelerate its deposition in the brain, which was consistent with the learning and memory impairment of AD mice, likely dependent on the membrane receptor of p75NTR. CONCLUSIONS: Our findings suggest that proBDNF may exert a crucially negative effect during AD pathogenesis andprogression.

Prothymosin Alpha: An Alarmin and More...

BACKGROUND/OBJECTIVE: Prothymosin alpha (proTα) is a ubiquitous polypeptide first isolated by Haritos in 1984, whose role still remains partly elusive. We know that proTα acts both, intracellularly, as an anti-apoptotic and proliferation mediator, and extracellularly, as a biologic response modifier mediating immune responses similarly to molecules termed as "alarmins". Our research team pioneered the elucidation of the mechanisms underlying the observed activities of proTα. RESULTS: We were the first to demonstrate that proTα levels increase during normal and abnormal cell proliferation. We showed that proTα acts pleiotropically, inducing immunomodulatory effects on immune cell populations. We revealed that the immunoreactive region of proTα is the carboxyterminal decapeptide proTα(100-109) and both molecules stimulate innate immune responses, signaling through Toll-like receptors (TLRs), specifically TLR-4. We reported that proTα and proTα(100-109) bind on the surface of human neutrophils on sites involving TLR-4, and cell activation is complemented by cytoplasmic calcium ion influx. Further, we showed that proTα and proTα(100-109) act as adjuvants upstream of lymphocyte stimulation and, in the presence of antigen, promote the expansion of antigen-reactive effectors. Most recently, we reported that proTα(100-109) may accumulate in experimentally inflamed sites and can serve as a surrogate biomarker in severe bacterial infections, proposing that extracellular release of proTα or proTα(100- 109) alerts the immune system during conditions of danger. CONCLUSION: We, therefore, suggest that proTα, and likely proTα(100-109), act as alarmins, being important immune mediators as well as biomarkers, and could eventually become targets for new therapeutic/diagnostic approaches in immune-related diseases like cancer, inflammation, and sepsis.

Antinociceptive Effect of Intrathecal Injection of Genetically Engineered Human Bone Marrow Stem Cells Expressing the Human Proenkephalin Gene in a Rat Model of Bone Cancer Pain.

Background. This study aimed to investigate the use of human bone marrow mesenchymal stem cells (hBMSCs) genetically engineered with the human proenkephalin (hPPE) gene to treat bone cancer pain (BCP) in a rat model. Methods. Primary cultured hBMSCs were passaged and modified with hPPE, and the cell suspensions (6 × 106) were then intrathecally injected into a rat model of BCP. Paw mechanical withdrawal threshold (PMWT) was measured before and after BCP. The effects of hPPE gene transfer on hBMSC bioactivity were analyzed in vitro and in vivo. Results. No changes were observed in the surface phenotypes and differentiation of hBMSCs after gene transfer. The hPPE-hBMSC group showed improved PMWT values on the ipsilateral side of rats with BCP from day 12 postoperatively, and the analgesic effect was reversed by naloxone. The levels of proinflammatory cytokines such as IL-1ß and IL-6 were ameliorated, and leucine-enkephalin (L-EK) secretion was augmented, in the hPPE-engineered hBMSC group. Conclusion. The intrathecal administration of BMSCs modified with the hPPE gene can effectively relieve pain caused by bone cancer in rats and might be a potentially therapeutic tool for cancer-related pain in humans.

Development of Precision Small-Molecule Proneurotrophic Therapies for Neurodegenerative Diseases.

Age-related neurodegenerative diseases, such as Alzheimer's disease, will represent one of the largest future burdens on worldwide healthcare systems due to the increasing proportion of elderly in our society. As deficiencies in neurotrophins are implicated in the pathogenesis of many age-related neurodegenerative disorders, it is reasonable to consider that global neurotrophin resistance may also become a major healthcare threat. Central nervous system networks are effectively maintained through aging by neuroprotective and neuroplasticity signaling mechanisms which are predominantly controlled by neurotrophin receptor signaling. Neurotrophin receptors are single pass receptor tyrosine kinases that form dimeric structures upon ligand binding to initiate cellular signaling events that control many protective and plasticity-related pathways. Declining functionality of the neurotrophin ligand-receptor system is considered one of the hallmarks of neuropathological aging. Therefore, it is imperative to develop effective therapeutic strategies to contend with this significant issue. While the therapeutic applications of cognate ligands for neurotrophin receptors are limited, the development of nonpeptidergic, small-molecule ligands can overcome these limitations, and productively regulate this important receptor system with beneficial effects. Using our advanced knowledge of the high-dimensionality complexity of receptor systems, the future generation of precision medicines targeting these systems will be an attainable goal.

Deletion of Nuclear Localizing Signal Attenuates Proinflammatory Activity of Prothymosin-Alpha and Enhances Its Neuroprotective Effect on Transient Ischemic Stroke.

Post-ischemic inflammation plays an important role in the progression of ischemia/reperfusion injuries. Prothymosin-α (ProT) can protect cells from necrotic death following ischemia; however, its immunostimulatory actions may counteract the neuroprotective effect. We proposed that ProTΔNLS, synthesized by deleting its nuclear localizing signal (NLS) at the C-terminal of ProT, can attenuate the immunostimulatory activity and has more salient neuroprotective effect. In this study, we examined the therapeutic effects of ProT and ProTΔNLS in a transient middle cerebral artery occlusion (tMCAO) model of rats. Rats that had sustained 90 min of tMCAO were treated with GST-vehicle, ProT, or ProTΔNLS. Therapeutic outcomes were evaluated by infarction volume assay and behavioral assessment. Changes to inflammatory mediators, including tumor necrosis factor α (TNF-α), interleukin-10 (IL-10), and myeloperoxidase (MPO) were evaluated by enzyme-linked immunosorbent assay. Activated matrix metalloproteinases 2 (MMP-2) and 9 (MMP-9) levels were evaluated by gelatin zymography. Microglial activation was identified by double-immunostaining for Iba-1 and CD68. Our results showed that while both ProT and ProTΔNLS reduce infarction volume and improve functional outcome, ProTΔNLS provides the best therapeutic outcome. ProT increases TNF-α but decreases IL-10 secretion after ischemic injury, reflecting its pro-inflammatory activity. ProTΔNLS suppresses expression of TNF-α, MPO, and activity of MMPs in ischemic brain tissue. It also suppresses activation of microglia in penumbral cortex. These data demonstrate the immunesuppressive activities of ProTΔNLS. In conclusion, ProT has pro-inflammatory effect that may counteract its neuroprotective effect. Deletion of NLS from ProT may attenuate post-ischemic inflammation and enhance the neuroprotective effects of ProT.

Local intramuscular injection of a plasmid encoding human proenkepahlin attenuates incision pain in rats.

We investigated the antinociceptive effect of local intramuscular injection of a plasmid encoding human proenkephalin (pVAX1-hPPE) on postoperative pain in rats. Male Sprague-Dawley rats with incision-induced pain were intramuscularly injected into injured plantaris muscle with empty vector (pVAX1) or pVAX1-hPPE, respectively. Paw mechanical threshold and thermal latency in the 200µg pVAX1-hPPE treated rats were significantly higher at 6h and on 1day, and lasted until day 7 after intramuscular administration, respectively. The analgesic effects were reversed by methylnaltrexone, suggesting that the antinociceptive effect of pVAX1-hPPE was mediated through peripheral opioid receptor pathway. In contrast, incisional or pVAX1-treated rats did not significantly affect pain thresholds. These results demonstrated that single intramuscular injection of pVAX1-hPPE attenuated incision-induced pain in rats, and it is worthy of further study as a potential gene therapy for postoperative pain.

Neuroprotective DAMPs member prothymosin alpha has additional beneficial actions against cerebral ischemia-induced vascular damages.

Prothymosin alpha (ProTα) suppresses stress-induced necrosis of cultured cortical neurons. As neuroprotection alone could not explain the long-lasting protective actions against cerebral ischemia by ProTα, we further examined whether ProTα, in addition to neuroprotective effects, has other anti-ischemic activities. When recombinant mouse ProTα (rmProTα) at 0.3 mg/kg was intravenously (i.v.) given 2 h after the start of tMCAO, all mice survived for more than 14 days. In evaluation of CD31- and tomato lectin-labeling as well as IgG and Evans blue leakage, rmProTα treatment (0.1 mg/kg) largely blocked ischemia-induced vascular damages. Therefore, rmProTα has novel beneficial effects against ischemia-induced brain damage through vascular mechanisms.

Prothymosin Alpha and Immune Responses: Are We Close to Potential Clinical Applications?

The thymus gland produces soluble molecules, which mediate significant immune functions. The first biologically active thymic extract was thymosin fraction V, the fractionation of which led to the isolation of a series of immunoactive polypeptides, including prothymosin alpha (proTα). ProTα displays a dual role, intracellularly as a survival and proliferation mediator and extracellularly as a biological response modifier. Accordingly, inside the cell, proTα is implicated in crucial intracellular circuits and may serve as a surrogate tumor biomarker, but when found outside the cell, it could be used as a therapeutic agent for treating immune system deficiencies. In fact, proTα possesses pleiotropic adjuvant activity and a series of immunomodulatory effects (eg, anticancer, antiviral, neuroprotective, cardioprotective). Moreover, several reports suggest that the variable activity of proTα might be exerted through different parts of the molecule. We first reported that the main immunoactive region of proTα is the carboxy-terminal decapeptide proTα(100-109). In conjunction with data from others, we also revealed that proTα and proTα(100-109) signal through Toll-like receptor 4. Although their precise molecular mechanism of action is yet not fully elucidated, proTα and proTα(100-109) are viewed as candidate adjuvants for cancer immunotherapy. Here, we present a historical overview on the discovery and isolation of thymosins with emphasis on proTα and data on some immune-related new activities of the polypeptide and smaller immunostimulatory peptides thereof. Finally, we propose a compiled scenario on proTα's mode of action, which could eventually contribute to its clinical application.

Higher diagnostic accuracy and cost-effectiveness using procalcitonin in the treatment of emergency medicine patients with fever (The HiTEMP study): a multicenter randomized study.

BACKGROUND: Fever is a common symptom in the emergency department(ED). Fever can be caused by bacterial infections, which are treated with antibiotics. Often, bacterial infections cannot be ruled out in the ED using standard diagnostics, and empiric antibiotic treatment is started. Procalcitonin(PCT) is a biomarker for bacterial infections, but its role in an undifferentiated ED population remains unclear. We hypothesize that PCT-guided therapy may reduce antibiotics prescription in undifferentiated febrile ED patients. The primary objectives of this study are to determine a) the efficacy, b) the safety of PCT-guided therapy, and c) the accuracy of the biomarker PCT for bacterial infections. The secondary objective is to study the cost-effectiveness of PCT-guided therapy. METHODS/DESIGN: This is a multicenter noninferiority randomized controlled trial. All adult ED patients with fever(≥38.2 °C) are randomized between standard care with and without the addition of a PCT level, after written informed consent. a) For efficacy, the reduction of patients receiving antibiotics is calculated, using a superiority analysis: differences between the PCT-guided group and control group are assessed using a Fisher's exact test, and a multivariable logistic regression analysis to account for the effects of demographic and medical variables on the percentage of febrile patients receiving antibiotics. b) Safety consists of a composite endpoint, defined as mortality, intensive care admission and ED return visit within 14 days. Noninferiority of PCT will be tested using a one-sided 95 % confidence interval for the difference in the composite safety endpoint between the PCT-guided and control groups using a noninferiority margin of 7.5 %. c) Accuracy of PCT and CRP for the diagnosis of bacterial infections will be reported, using the sensitivity, specificity, and the area under the receiver-operating-characteristic curve in the definitive diagnosis of bacterial infections. The sample size is 550 patients, which was calculated using a power analysis for all primary objectives. Enrollment of patients started in August 2014 and will last 2 years. DISCUSSION: PCT may offer a more tailor-made treatment to the individual ED patient with fever. Prospective costs analyses will reveal the economic consequences of implementing PCT-guided therapy in the ED. THIS TRIAL IS REGISTERED IN THE DUTCH TRIAL REGISTER: NTR4949.

Provider Decisions to Treat Respiratory Illnesses with Antibiotics: Insights from a Randomized Controlled Trial.

RATIONALE: Lower respiratory tract illness (LRTI) frequently causes adult hospitalization and antibiotic overuse. Procalcitonin (PCT) treatment algorithms have been used successfully in Europe to safely reduce antibiotic use for LRTI but have not been adopted in the United States. We recently performed a feasibility study for a randomized clinical trial (RCT) of PCT and viral testing to guide therapy for non-pneumonic LRTI. OBJECTIVE: The primary objective of the current study was to understand factors influencing PCT algorithm adherence during the RCT and evaluate factors influencing provider antibiotic prescribing practices for LRTI. STUDY DESIGN: From October 2013-April 2014, 300 patients hospitalized at a community teaching hospital with non-pneumonic LRTI were randomized to standard or PCT-guided care with viral PCR testing. Algorithm adherence data was collected and multivariate stepwise logistic regression of clinical variables used to model algorithm compliance. 134 providers were surveyed anonymously before and after the trial to assess knowledge of biomarkers and viral testing and antibiotic prescribing practices. RESULTS: Diagnosis of pneumonia on admission was the only variable significantly associated with non-adherence [7% (adherence) vs. 26% (nonadherence), p = 0.01]. Surveys confirmed possible infiltrate on chest radiograph as important for provider decisions, as were severity of illness, positive sputum culture, abnormal CBC and fever. However, age, patient expectations and medical-legal concerns were also at least somewhat important to prescribing practices. Physician agreement with the importance of viral and PCT testing increased from 42% to 64% (p = 0.007) and 49% to 74% (p = 0.001), respectively, after the study. CONCLUSIONS: Optimal algorithm adherence will be important for definitive PCT intervention trials in the US to determine if PCT guided algorithms result in better outcomes than reliance on traditional clinical variables. Factors influencing treatment decisions such as patient age, presence of fever, patient expectations and medical legal concerns may be amenable to education to improve PCT algorithm compliance for LRTI.

Hormonal control of T-cell development in health and disease.

The physiology of the thymus, the primary lymphoid organ in which T cells are generated, is controlled by hormones. Data from animal models indicate that several peptide and nonpeptide hormones act pleiotropically within the thymus to modulate the proliferation, differentiation, migration and death by apoptosis of developing thymocytes. For example, growth hormone and prolactin can enhance thymocyte proliferation and migration, whereas glucocorticoids lead to the apoptosis of these developing cells. The thymus undergoes progressive age-dependent atrophy with a loss of cells being generated and exported, therefore, hormone-based therapies are being developed as an alternative strategy to rejuvenate the organ, as well as to augment thymocyte proliferation and the export of mature T cells to peripheral lymphoid organs. Some hormones (such as growth hormone and progonadoliberin-1) are also being used as therapeutic agents to treat immunodeficiency disorders associated with thymic atrophy, such as HIV infection. In this Review, we discuss the accumulating data that shows the thymus gland is under complex and multifaceted hormonal control that affects the process of T-cell development in health and disease.

Antimicrobial stewardship in the intensive care setting--a review and critical appraisal of the literature.

BACKGROUND: Many antimicrobial stewardship programmes (ASPs) target the intensive care unit owing to high antimicrobial utilisation. In this review, we summarise and assess the quality of evidence supporting the implementation of various ASP strategies in the intensive care unit setting with a focus on publications between 2010 and 2015. METHODS: We searched Medline up to April 2015 and screened publications of interest for additional relevant articles. We grouped the strategies into four categories: audit and feedback, formulary restrictions, guidelines/clinical pathways, and procalcitonin. We used GRADE terminology to describe the quality of evidence. RESULTS AND CONCLUSIONS: We identified several studies reporting optimisation and reduction of antibiotic utilisation as well as cost reduction in all four strategies. Randomised controlled trials reviewing the role of procalcitonin demonstrate a moderate level of evidence. Given the lack of randomised controlled trials to support the role of guidelines, formulary restrictions, and audit and feedback, the level of evidence supporting these strategies is low. Importantly, there is no convincing evidence to support the main goal of ASP, namely to improve patient outcomes. Larger, rigorous long-term studies using a cluster randomised controlled trial or at least a controlled quasi-experimental design with time series are required to assess the impact of ASP on patient-important outcomes and on the emergence of resistance in the intensive care unit setting.

Functional Characterization of Human ProNGF and NGF Mutants: Identification of NGF P61SR100E as a "Painless" Lead Investigational Candidate for Therapeutic Applications.

BACKGROUND: Nerve Growth Factor (NGF) holds a great therapeutic promise for Alzheimer's disease, diabetic neuropathies, ophthalmic diseases, dermatological ulcers. However, the necessity for systemic delivery has hampered the clinical applications of NGF due to its potent pro-nociceptive action. A "painless" human NGF (hNGF R100E) mutant has been engineered. It has equal neurotrophic potency to hNGF but a lower nociceptive activity. We previously described and characterized the neurotrophic and nociceptive properties also of the hNGF P61S and P61SR100E mutants, selectively detectable against wild type hNGF. However, the reduced pain-sensitizing potency of the "painless" hNGF mutants has not been quantified. OBJECTIVES AND RESULTS: Aiming at the therapeutic application of the "painless" hNGF mutants, we report on the comparative functional characterization of the precursor and mature forms of the mutants hNGF R100E and hNGF P61SR100E as therapeutic candidates, also in comparison to wild type hNGF and to hNGF P61S. The mutants were assessed by a number of biochemical, biophysical methods and assayed by cellular assays. Moreover, a highly sensitive ELISA for the detection of the P61S-tagged mutants in biological samples has been developed. Finally, we explored the pro-nociceptive effects elicited by hNGF mutants in vivo, demonstrating an expanded therapeutic window with a ten-fold increase in potency. CONCLUSIONS: This structure-activity relationship study has led to validate the concept of developing painless NGF as a therapeutic, targeting the NGF receptor system and supporting the choice of hNGF P61S R100E as the best candidate to advance in clinical development. Moreover, this study contributes to the identification of the molecular determinants modulating the properties of the hNGF "painless" mutants.