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The relative concentrations of antibodies produced in monkeys against three forms of human growth hormone (hGH) were determined using an antigen-specific avidin/biotin ELISA assay. Monkeys were treated in two separate 90-day studies with recombinant methionyl-hGH (met-hGH) and pituitary-derived hGH (pit-hGH) (Study 1) and recombinant natural sequence hGH (Study 2). The lowest dose was equal to the expected therapeutic dose of 0.1 IU/kg. Sixty-nine percent of monkeys treated with pit-hGH and 81% of those treated with met-hGH developed detectable anti-hGH responses. The magnitudes of the responses exhibited wide animal to animal variability, were not markedly related to dose or sex, and were lower than levels obtained in monkeys immunized with hGH in Freund's adjuvant. In contrast, the incidence of antibody responses in monkeys treated with natural sequence hGH was lower (23% in one experiment and 5% in a replicate experiment) and took longer to develop. Antibody concentrations were lower, on average, than in those animals treated with met- or pit-hGH. These results are in accord with those observed clinically, thus supporting the use of the monkey model to predict the relative immunogenicity of some proteins in humans. [ABSTRACT FROM PUBLISHER]

Background: Dual antiplatelet therapy (DAPT) is commonly employed for neuroendovascular stenting due to the significant risk of thromboembolism. Clopidogrel and aspirin are most often selected as initial DAPTs; however, there is limited literature available to support guidance of DAPT in this setting. The objective of this study was to evaluate safety and efficacy in patients whose final regimen included either DAPT with aspirin and clopidogrel (DAPT-C) or DAPT with aspirin and ticagrelor (DAPT-T)., Methods: This was a multicenter, retrospective cohort of patients who underwent neuroendovascular stenting and received DAPT between July 1, 2017, and October 31, 2020. Study participants were allocated into groups based on discharge DAPT regimen. The primary outcome was incidence of stent thrombosis at 3-6 months on DAPT-C versus DAPT-T, as defined by the presence of thrombus on imaging or new onset stroke. Secondary outcomes included major and minor bleeding and death within 3-6 months after the procedure., Results: Five hundred and seventy patients were screened across 12 sites. Of those, 486 were included (DAPT-C n = 360, DAPT-T n = 126). There was no difference in the primary outcome of stent thrombosis between the DAPT-C and DAPT-T groups (8% vs. 8%, p = 0.97) and no difference in any of the secondary safety outcomes., Conclusions: Using DAPT-C or DAPT-T regimens in a broad population of neuroendovascular stenting procedures appears to have similar safety and efficacy profiles. Further prospective evaluation is warranted to streamline the practice of DAPT selection and monitoring to determine the impact on clinical outcomes., (© 2023. Springer Science+Business Media, LLC, part of Springer Nature and Neurocritical Care Society.)

Background: Emergent neuroendovascular stenting presents challenges for the utilization of antiplatelet agents., Methods: This was a multicenter, retrospective cohort of patients who underwent emergent neuroendovascular stenting. The primary endpoints were thrombotic and bleeding events in relation to the timing of antiplatelet administration, route of administration, and choice of intravenous (IV) agent and the study investigated practice variability in antiplatelet utilization., Results: Five-hundred and seventy patients were screened across 12 sites. Of those, 167 were included for data analysis. For patients who presented with ischemic stroke, artery dissection and emergent internal carotid artery (ICA) stenting who received an antiplatelet agent prior to or during the procedure, 57% were given an IV antiplatelet agent; for patients who were given an antiplatelet agent after the procedure, 96% were given an oral agent. For patients who presented for aneurysm repair and received an antiplatelet agent prior to or during the procedure, 74% were given an IV agent; patients who were given an antiplatelet agent after the completion of the procedure were given an oral antiplatelet agent 90% of the time. In patients who presented with ischemic stroke, artery dissection and emergent ICA stenting who received oral antiplatelet agents post-procedure were more likely to have thrombotic events compared to those who received oral antiplatelet agents prior to or during the procedure (29% vs 9%; p   =  0.04). There were no differences in the primary outcomes observed when comparing other antiplatelet treatment strategies., Conclusion: The optimal timing of antiplatelet administration in relation to stent placement and route of administration of antiplatelet agents is unclear. Timing and route of administration of antiplatelet agents may have an effect on thrombosis in emergent neuroendovascular stenting. Significant practice variation exists in antiplatelet agent utilization in emergent neuroendovascular stenting.

Adeno-associated virus (AAV) has emerged as a leading platform for gene delivery for treating various diseases due to its excellent safety profile and efficient transduction to various target tissues. However, the large-scale production and long-term storage of viral vectors is not efficient resulting in lower yields, moderate purity, and shorter shelf-life compared to recombinant protein therapeutics. This review provides a comprehensive analysis of upstream, downstream and formulation unit operation challenges encountered during AAV vector manufacturing, and discusses how desired product quality attributes can be maintained throughout product shelf-life by understanding the degradation mechanisms and formulation strategies. The mechanisms of various physical and chemical instabilities that the viral vector may encounter during its production and shelf-life because of various stressed conditions such as thermal, shear, freeze-thaw, and light exposure are highlighted. The role of buffer, pH, excipients, and impurities on the stability of viral vectors is also discussed. As such, the aim of this review is to outline the tools and a potential roadmap for improving the quality of AAV-based drug products by stressing the need for a mechanistic understanding of the involved processes., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Little data exist regarding the practice of sodium management in acute neurologically injured patients. This study describes the practice variations, thresholds for treatment, and effectiveness of treatment in this population., Methods: This retrospective, multicenter, observational study identified 400 ICU patients, from 17 centers, admitted for ≥48 h with subarachnoid hemorrhage (SAH), traumatic brain injury (TBI), intraparenchymal hemorrhage, or intracranial tumors between January 1, 2011 and July 31, 2012. Data collection included demographics, APACHE II, Glascow Coma Score (GCS), serum sodium (Na+), fluid rate and tonicity, use of sodium-altering therapies, intensive care unit (ICU) and hospital length of stay, and modified Rankin score upon discharge. Data were collected for the first 21 days of ICU admission or ICU discharge, whichever came first. Sodium trigger for treatment defined as the Na+ value prior to treatment with response defined as an increase of ≥4 mEq/L at 24 h., Results: Sodium-altering therapy was initiated in 34 % (137/400) of patients with 23 % (32/137) having Na + >135 mEq/L at time of treatment initiation. The most common indications for treatment were declining serum Na + (68/116, 59 %) and cerebral edema with mental status changes (21/116, 18 %). Median Na + treatment trigger was 133 mEq/L (IQR 129-139) with no difference between diagnoses. Incidence and treatment of hyponatremia was more common in SAH and TBI [SAH (49/106, 46 %), TBI (39/97, 40 %), ICH (27/102, 26 %), tumor (22/95, 23 %); p = 0.001]. The most common initial treatment was hypertonic saline (85/137, 62 %), followed by oral sodium chloride tablets (42/137, 31 %) and fluid restriction (15/137, 11 %). Among treated patients, 60 % had a response at 24 h. Treated patients had lower admission GCS (12 vs. 14, p = 0.02) and higher APACHE II scores (12 vs. 10, p = 0.001). There was no statistically significant difference in outcome when comparing treated and untreated patients., Conclusion: Sodium-altering therapy is commonly employed among neurologically injured patients. Hypertonic saline infusions were used first line in more than half of treated patients with the majority having a positive response at 24 h. Further studies are needed to evaluate the impact of various treatments on patient outcomes.

Emergency neurological life support (ENLS) is an educational program designed to provide users advisory instruction regarding management for the first few hours of a neurological emergency. The content of the course is divided into 14 modules, each addressing a distinct category of neurological injury. The course is appropriate for practitioners and providers from various backgrounds who work in environments of variable medical complexity. The focus of ENLS is centered on a standardized treatment algorithm, checklists to guide early patient care, and a structured format for communication of findings and concerns to other healthcare professionals. Certification and training in ENLS is hosted by the Neurocritical Care Society. This document introduces the concept of ENLS and describes the revisions that constitute this second version.

Careful patient monitoring using a variety of techniques including clinical and laboratory evaluation, bedside physiological monitoring with continuous or non-continuous techniques and imaging is fundamental to the care of patients who require neurocritical care. How best to perform and use bedside monitoring is still being elucidated. To create a basic platform for care and a foundation for further research the Neurocritical Care Society in collaboration with the European Society of Intensive Care Medicine, the Society for Critical Care Medicine and the Latin America Brain Injury Consortium organized an international, multidisciplinary consensus conference to develop recommendations about physiologic bedside monitoring. This supplement contains a Consensus Summary Statement with recommendations and individual topic reviews as a background to the recommendations. In this article, we highlight the recommendations and provide additional conclusions as an aid to the reader and to facilitate bedside care.

A variety of technologies have been developed to assist decision-making during the management of patients with acute brain injury who require intensive care. A large body of research has been generated describing these various technologies. The Neurocritical Care Society (NCS) in collaboration with the European Society of Intensive Care Medicine (ESICM), the Society for Critical Care Medicine (SCCM), and the Latin America Brain Injury Consortium (LABIC) organized an international, multidisciplinary consensus conference to perform a systematic review of the published literature to help develop evidence-based practice recommendations on bedside physiologic monitoring. This supplement contains a Consensus Summary Statement with recommendations and individual topic reviews on physiologic processes important in the care of acute brain injury. In this article we provide the evidentiary tables for select topics including systemic hemodynamics, intracranial pressure, brain and systemic oxygenation, EEG, brain metabolism, biomarkers, processes of care and monitoring in emerging economies to provide the clinician ready access to evidence that supports recommendations about neuromonitoring.

Objective: Most state Medicaid programs reimburse physicians for providing fluoride varnish, yet the only published studies of cost-effectiveness do not show cost-savings. Our objective is to apply state-specific claims data to an existing published model to quickly and inexpensively estimate the cost-savings of a policy consideration to better inform decisions - specifically, to assess whether Indiana Medicaid children's restorative service rates met the threshold to generate cost-savings., Methods: Threshold analysis was based on the 2006 model by Quiñonez et al. Simple calculations were used to "align" the Indiana Medicaid data with the published model. Quarterly likelihoods that a child would receive treatment for caries were annualized. The probability of a tooth developing a cavitated lesion was multiplied by the probability of using restorative services. Finally, this rate of restorative services given cavitation was multiplied by 1.5 to generate the threshold to attain cost-savings. Restorative services utilization rates, extrapolated from available Indiana Medicaid claims, were compared with these thresholds., Results: For children 1-2 years old, restorative services utilization was 2.6 percent, which was below the 5.8 percent threshold for cost-savings. However, for children 3-5 years of age, restorative services utilization was 23.3 percent, exceeding the 14.5 percent threshold that suggests cost-savings., Conclusions: Combining a published model with state-specific data, we were able to quickly and inexpensively demonstrate that restorative service utilization rates for children 36 months and older in Indiana are high enough that fluoride varnish regularly applied by physicians to children starting at 9 months of age could save Medicaid funds over a 3-year horizon., (© 2013 American Association of Public Health Dentistry.)

Background: Numerous anticonvulsant agents are now available for treating status epilepticus (SE). However, a paucity of data is available to guide clinicians in the initial treatment of seizures or SE. This study describes the current strategies being employed to treat SE in the U.S.A., Methods: Fifteen American academic medical centers completed a retrospective, multicenter, observational study by reviewing 10-20 of the most recent cases of SE at their institution prior to December 31, 2009. A multivariate analysis was performed to determine factors associated with cessation of seizures., Results: A total of 150 patients were included. Most patients with SE had a seizure disorder (58%). SE patients required a median of 3 AEDs for treatment. Three quarters of patients received a benzodiazepine as first-line therapy (74.7%). Phenytoin (33.3%) and levetiracetam (10%) were commonly used as the second AED. Continuous infusions of propofol, barbiturate, or benzodiazepine were used in 36% of patients. Median time to resolution of SE was 1 day and was positively associated with presence of a complex partial seizure, AED non-compliance prior to admission, and lorazepam plus another AED as initial therapy. Prolonged ICU length of stay and topiramate therapy prior to admission were negatively associated with SE resolution. Mortality was higher in patients without a history of seizure (22.2 vs. 6.9%, p = 0.006)., Conclusions: The use of a benzodiazepine followed by an AED, such as phenytoin or levetiracetam, is common as first and second-line therapy for SE and appears to be associated with a shorter time to SE resolution. AED selection thereafter is highly variable. Patients without a history of seizure who develop SE had a higher mortality rate.

Background: Advances in the understanding of human biochemistry and physiology have provided insight into new pathways by which we can understand traumatic brain injury (TBI). Increased sophistication of laboratory techniques and developments in the field of proteomics has led to the discovery and rapid detection of new biomarkers not previously available., Objective: To review recent advances in biomarker research for traumatic brain injury, describe the features of the ideal biomarker and to explore the potential role of these biomarkers in improving clinical management of brain injured patients., Methods: Through a literature review of recent research on TBI biomarkers and through experience with TBI research, important elements of biomarker development are described together with potential applications to patient care., Conclusions: TBI biomarkers could have a significant impact on patient care by assisting in the diagnosis, risk stratification and management of TBI. Biomarkers could provide major opportunities for the conduct of clinical research, including confirmation of injury mechanism(s) and drug target identification. Continuing studies by the authors' group are now being conducted to elucidate more fully the relationships between new biomarkers and severity of injury and clinical outcomes in all severities of TBI patients.

Objective: To review the published clinical data assessing the role of adjunctive rifampin therapy for the treatment of staphylococcal central nervous system (CNS) infections., Data Sources: A MEDLINE search (January 1966-March 2000) of English-language literature pertaining to CNS staphylococcal infections and rifampin was performed; tertiary sources were also used., Data Extraction: Human data and case reports were included, as no clinical trials have been published., Data Synthesis: Retrospective reviews of rifampin used in combination with other antibiotics for serious bacterial infections show conflicting results. Few case reports have described clinical successes with adjunctive rifampin therapy for CNS infections., Conclusions: The routine use of adjunctive rifampin for CNS infections cannot be justified.

Management of sedation and analgesia in critical care medicine is a multidisciplinary process that involves physicians, nurses, pharmacists, and other healthcare providers. Optimal management of these common issues includes recognition of the importance of predisposing and causative conditions that contribute to the sensations of pain and discomfort, anxiety, and delirium. Treatment includes pharmacological intervention, correction of predisposing factors, and use of other preventative and nonpharmacological measures. It is increasingly clear that, although necessary for patient comfort, sedative and analgesic medications can have adverse consequences, including side-effects as well as prolonged mechanical ventilation and ICU length of stay. Optimal use of sedative and analgesic medications involves matching unique properties of specific medications with individual patient characteristics. Guidelines that minimize unnecessary variability in practice, prevent excessive medication, and emphasize management based on individual patient characteristics improve the effective utilization of these medications.

p56(lck) is a lymphocyte-specific tyrosine kinase that plays an important role in both T-cell maturation and activation. We have developed a homogeneous assay in which p56(lck) catalyzes the transfer of the gamma-phosphate group from [gamma-(33)P]ATP to a biotinylated peptide substrate. The labeled peptide is then captured on a streptavidin-coated scintillation proximity assay (SPA) bead or imaging proximity bead. The SPA is counted in a microplate scintillation counter and the imaging proximity assay is counted in a charge-coupled device-based imaging system called LEADseekertrade mark, recently launched as a homogeneous imaging system by Amersham Pharmacia Biotech. We show, via time-dependence assays and inhibitor studies, that this assay can be performed in 1536-well microplate format using imaging proximity as the method of detection. The results compare favorably with the same assay performed in 384-well microplate format using both SPA and imaging proximity as the detection methods. From this study, we conclude that a kinase assay can be performed in 384- and 1536-well format using imaging as the detection method, with significant time savings over standard scintillation counting. In addition, we show cost saving advantages of 1536- over 384-well format in terms of reagent usage, higher throughput, and waste disposal.

The translocation of phospholipids across the plasma membrane has been widely documented as one of the earliest measurable biochemical events of apoptosis. Using fluorescently labelled annexin V, which preferentially binds phosphatidylserine (PS) in the presence of Ca(2+), the externalization of PS can be measured and apoptosis quantified using flow cytometry. Conventional detection methods utilizing annexin V, while faster than in situ DNA end-labelling or DNA laddering, require extensive sample preparation which may compromise samples and makes rapid, high volume screening prohibitive. This paper describes a novel assay for the measurement of apoptosis based upon binding of radiolabelled annexin V to apoptotic cells attached to the growth surface of a 96-well scintillating microplate (Cytostar-T(R)). We compared measurements of apoptosis made by flow cytometry to those obtained with the scintillating microplate in three model systems, treatment of: mouse connective tissue (L-M) cells with lymphotoxin (LT), human lung carcinoma (H460) cells with Apo-2 ligand and human umbilical vein endothelial (HUVE) cells with staurosporine. In this assay, we compare both direct and indirect labelling methods by utilizing either iodinated annexin V or biotinylated annexin V/[(35)S] streptavidin to radiolabel apoptotic cells. The signal detected is a direct consequence of the binding of annexin V to externalized PS on apoptotic cells and the proximity of the label to the base of the plate. Using this method, separation of bound and unbound radiolabel signal occurs directly within the well resulting in a sensitive assay that requires minimal manipulation and can accomodate a large number of samples.

A 5-year study of graduates' performance on NCLEX-RN was conducted using data from July 1988 through February 1994. This time frame related to the "new" test plan introduced in 1988 with pass/fail results for NCLEX analysis. Using a quota sampling technique of 188 graduates, selected admission and curriculum variables and National League for Nursing (NLN) Comprehensive Achievement Test scores were studied in relationship to NCLEX-RN examination results. The strongest indicators of success were SAT verbal scores, nursing grade point average, and NLN Comprehensive Achievement Test scores. In addition, logistic regression analyses identified three nursing courses in combination with the NLN Comprehensive Achievement Test score as a strong model for prediction. Even though pass/fail data limit statistical analyses, the predictor variables were strong at P = 0.0001. These findings are consistent with prior studies.

[14C]Thymidine uptake into V-79 hamster lung fibroblasts has been successfully demonstrated using a noninvasive, real-time method utilizing Cytostar-T scintillating microplates. These plates are standard format, tissue culture-treated, 96-well microplates with an integral scintillating base. The microplates permit the culture and observation of adherent cell monolayers. Biological activities of the cells can be studied by the provision of specific radiolabeled compounds. The biological activities of the adherent monolayer bring the specific radiolabel into proximity with the scintillating base and a scintillation signal is thereby generated. [14C]Thymidine incorporation on the microplates can be used to examine cell proliferation and cell cycle events. Using a combined mitotic shake-off/aphidicolin treatment to achieve synchronization, the thymidine incorporation activities of V-79 cells have been examined on Cytostar-T plates and correlated to traditional methods of determining incorporation. The method was further used to examine the effects of colcemid and olomoucine, both chemical inhibitors of cell proliferation, on synchronous populations of cells. The homogeneous detection format and the microplate nature of the method suggest a role for scintillating microplates in cell biology research and drug discovery.

Amiodarone is useful for the treatment of ventricular arrhythmias but has been associated with a significant degree of toxicity especially to lung and liver. The drug produces phospholipid accumulation in multilamella lysosomal inclusions in many tissues due to the ability of amiodarone and desethylamiodarone to inhibit phospholipase A. The adverse effects do not correlate with the plasma levels of amiodarone but relate more closely to the cumulative dose. No clear way of predicting amiodarone toxicity has yet emerged. In this report, normal human lymphoblasts in tissue culture were shown by electron microscopy to have dose-dependent increases in multilamellar inclusions when grown with amiodarone at concentrations which are routinely observed in patients receiving the drug. The content of phospholipid also increased but this parameter was not as sensitive as the number of multilamellar inclusions. Finally, lymphocytes from patients treated with amiodarone were examined by electron microscopy and shown to have increased numbers of multilamellar bodies.

The cytotoxic activity of chlorambucil as a function of pH was investigated in P388 tumor cells growing in static suspension culture. A decrease in extracellular pH from 7.8 to 7.2 was associated with a decrease in intracellular pH from 7.92 to 7.55. The cytotoxic potency of chlorambucil increased as the extracellular pH decreased; IC99 values were 20 and 60 microM when the extracellular pH was 7.2 and 7.8 respectively. Covalent binding to cellular macromolecules was about 1.9 times greater at pH 7.2 relative to that at pH 7.8. These results suggest that pH may be an important determinant of the oncotoxic specificity of chlorambucil, and that the cytotoxic activity of this agent could be selectively directed toward tumor cells by the selective manipulation of intracellular and extracellular pH. A potential influence of intracellular and extracellular pH on cytotoxic, mutagenic, carcinogenic, and teratogenic potencies of other chemicals is also suggested. Additionally, these investigations demonstrate the importance of carefully controlling pH throughout the drug exposure period when evaluating the relative potency of potential cytotoxic, mutagenic, carcinogenic, and teratogenic agents in cell or organ culture.

Walker 256 rat carcinosarcoma cells growing as solid subcutaneous or intramuscular tumors depressed hepatic microsomal mixed-function oxygenase activity to less than 20% of control activity, but the same tumor cells growing as free ascites cells in the peritoneal cavity did not. Necrosis of the core area of solid tumors was observed. Tumor cells may release a substance that depresses hepatic microsomal mixed-function oxygenase activity only upon their death.