Evaluating the Impact of Telehealth on Carbon Footprint During Three Phases of the Pandemic at a Rural Academic Medical Center.

Background: Climate change is primarily driven by greenhouse gases, such as carbon dioxide (CO2). Telehealth visits have been found to mitigate carbon emissions by reducing patient and physician transport. Dartmouth Hitchcock Medical Center (DHMC) is the most rural academic medical center in the country, serving a population where the majority of patients reach the hospital by car. No large study or systematic review has evaluated the impact of telehealth visits on CO2 emissions (CO2e) across multiple specialties in a purely rural setting. Further, no sizable rurally focused study has compared CO2e avoided during the various stages of the pandemic. Methods: We extracted data for all outpatient telehealth visits at DHMC from three periods: prepandemic, early pandemic, and late pandemic. The extracted data included the pandemic stage of the virtual visit, the type of visit (video or telephone), the specialty, and the distance from the patient's home to DHMC. Results: The total CO2e avoided among all three pandemic stages analyzed in this study was 23,658,898 kg (n = 251,832). During period 1, the mean driving distance = 159.0 miles; CO2e avoided per encounter = 128.3 kg; period 2, mean distance = 84.85 miles; average CO2e avoided per encounter = 68.47 CO2e kg; and period 3, mean distance = 112.9 miles; average CO2e avoided per encounter = 91.08 kg. Conclusions: This data supported long distances to the medical center and large savings in CO2e avoided across multiple specialties that spanned all pandemic periods. Further, this level of averted emissions could translate to over $3M in saved fuel costs and the avoidance of six excess deaths. While discussions of the future of telehealth commonly focus on access, use cases, technology, costs, and satisfaction, the impact on carbon footprint is an additional important metric, particularly in largely rural regions.

Outpatient Virtual Visits and the "Right" Amount of Telehealth Going Forward.

Background: An exponential increase in outpatient telehealth visits occurred early in the pandemic period that has been followed by volumes that, although lower than peak numbers, are substantially greater than the pre-pandemic period. This provided an opportunity to assess provider perceptions regarding the right prevalence going forward and key obstacles to achieving it. Methods: A 10-question survey was distributed to all outpatient providers within the Dartmouth-Hitchcock Health System. Domains included practice location, specialty, professional degree, experience with telehealth, satisfaction, perception of the amount of telehealth that could be adequately delivered going forward, role of audio-only, and obstacles. Results: Three hundred thirty-six providers completed the survey representing 51 specialties. The most common response regarding the proportion of outpatient visits that could be delivered by video going forward was 21-50% (n = 104) followed by 6-20% (n = 99) and >50% (n = 71). A minority of respondents chose ≤5% (n = 17). In terms of the fraction of video visits for which phone was equally effective, a similar percentage of respondents felt that it was 1/10 (22%), 1/4 (20%), or 1/2 (26%) of visits. Fewer felt that all (7%) or 3/4 (15%) of visits were equally effective, and 10% felt that it was none. Common obstacles identified were the need for a physical exam, unique aspects of providers' patients, patient preference, and issues regarding technology and internet speed/connectivity. Conclusions: After a period of exponential growth in virtual visits due to the pandemic, outpatient providers within an academic health system felt that a substantial portion of future visits could be delivered by this modality.

Telehealth's New Horizon: Providing Smart Hospital-Level Care in the Home.

During the COVID-19 pandemic, medical providers have expanded telehealth into daily practice, with many medical and behavioral health care visits provided remotely over video or through phone. The telehealth market was already facilitating home health care with increasing levels of sophistication before COVID-19. Among the emerging telehealth practices, telephysical therapy; teleneurology; telemental health; chronic care management of congestive heart failure, chronic obstructive pulmonary disease, diabetes; home hospice; home mechanical ventilation; and home dialysis are some of the most prominent. Home telehealth helps streamline hospital/clinic operations and ensure the safety of health care workers and patients. The authors recommend that we expand home telehealth to a comprehensive delivery of medical care across a distributed network of hospitals and homes, linking patients to health care workers through the Internet of Medical Things using in-home equipment, including smart medical monitoring devices to create a "medical smart home." This expanded telehealth capability will help doctors care for patients flexibly, remotely, and safely as a part of standard operations and during emergencies such as a pandemic. This model of "telehomecare" is already being implemented, as shown herein with examples. The authors envision a future in which providers and hospitals transition medical care delivery to the home just as, during the COVID-19 pandemic, students adapted to distance learning and adults transitioned to remote work from home. Many of our homes in the future may have a "smart medical suite" as well as a "smart home office."

The WNT-controlled transcriptional regulator LBH is required for mammary stem cell expansion and maintenance of the basal lineage.

The identification of multipotent mammary stem cells (MaSCs) has provided an explanation for the unique regenerative capacity of the mammary gland throughout adult life. However, it remains unclear what genes maintain MaSCs and control their specification into the two epithelial lineages: luminal and basal. LBH is a novel transcription co-factor in the WNT pathway with hitherto unknown physiological function. LBH is expressed during mammary gland development and aberrantly overexpressed in aggressive 'basal' subtype breast cancers. Here, we have explored the in vivo role of LBH in mammopoiesis. We show that in postnatal mammary epithelia, LBH is predominantly expressed in the Lin(-)CD29(high)CD24(+) basal MaSC population. Upon conditional inactivation of LBH, mice exhibit pronounced delays in mammary tissue expansion during puberty and pregnancy, accompanied by increased luminal differentiation at the expense of basal lineage specification. These defects could be traced to a severe reduction in the frequency and self-renewal/differentiation potential of basal MaSCs. Mechanistically, LBH induces expression of key epithelial stem cell transcription factor ΔNp63 to promote a basal MaSC state and repress luminal differentiation genes, mainly that encoding estrogen receptor α (Esr1/ERα). Collectively, these studies identify LBH as an essential regulator of basal MaSC expansion/maintenance, raising important implications for its potential role in breast cancer pathogenesis.

Increased toll-like receptor 4 in cerebral endothelial cells contributes to the astrocyte swelling and brain edema in acute hepatic encephalopathy.

Astrocyte swelling and the subsequent increase in intracranial pressure and brain herniation are major clinical consequences in patients with acute hepatic encephalopathy. We recently reported that conditioned media from brain endothelial cells (ECs) exposed to ammonia, a mixture of cytokines (CKs) or lipopolysaccharide (LPS), when added to astrocytes caused cell swelling. In this study, we investigated the possibility that ammonia and inflammatory agents activate the toll-like receptor 4 (TLR4) in ECs, resulting in the release of factors that ultimately cause astrocyte swelling. We found a significant increase in TLR4 protein expression when ECs were exposed to ammonia, CKs or LPS alone, while exposure of ECs to a combination of these agents potentiate such effects. In addition, astrocytes exposed to conditioned media from TLR4-silenced ECs that were treated with ammonia, CKs or LPS, resulted in a significant reduction in astrocyte swelling. TLR4 protein up-regulation was also detected in rat brain ECs after treatment with the liver toxin thioacetamide, and that thioacetamide-treated TLR4 knock-out mice exhibited a reduction in brain edema. These studies strongly suggest that ECs significantly contribute to the astrocyte swelling/brain edema in acute hepatic encephalopathy, likely as a consequence of increased TLR4 protein expression by blood-borne noxious agents.

Decreased astrocytic thrombospondin-1 secretion after chronic ammonia treatment reduces the level of synaptic proteins: in vitro and in vivo studies.

Chronic hepatic encephalopathy (CHE) is a major complication in patients with severe liver disease. Elevated blood and brain ammonia levels have been implicated in its pathogenesis, and astrocytes are the principal neural cells involved in this disorder. Since defective synthesis and release of astrocytic factors have been shown to impair synaptic integrity in other neurological conditions, we examined whether thrombospondin-1 (TSP-1), an astrocytic factor involved in the maintenance of synaptic integrity, is also altered in CHE. Cultured astrocytes were exposed to ammonia (NH4Cl, 0.5-2.5 mM) for 1-10 days, and TSP-1 content was measured in cell extracts and culture media. Astrocytes exposed to ammonia exhibited a reduction in intra- and extracellular TSP-1 levels. Exposure of cultured neurons to conditioned media from ammonia-treated astrocytes showed a decrease in synaptophysin, PSD95, and synaptotagmin levels. Conditioned media from TSP-1 over-expressing astrocytes that were treated with ammonia, when added to cultured neurons, reversed the decline in synaptic proteins. Recombinant TSP-1 similarly reversed the decrease in synaptic proteins. Metformin, an agent known to increase TSP-1 synthesis in other cell types, also reversed the ammonia-induced TSP-1 reduction. Likewise, we found a significant decline in TSP-1 level in cortical astrocytes, as well as a reduction in synaptophysin content in vivo in a rat model of CHE. These findings suggest that TSP-1 may represent an important therapeutic target for CHE. Defective release of astrocytic factors may impair synaptic integrity in chronic hepatic encephalopathy. We found a reduction in the release of the astrocytic matricellular proteins thrombospondin-1 (TSP-1) in ammonia-treated astrocytes; such reduction was associated with a decrease in synaptic proteins caused by conditioned media from ammonia-treated astrocytes. Exposure of neurons to CM from ammonia-treated astrocytes, in which TSP-1 is over-expressed, reversed (by approx 75%) the reduction in synaptic proteins. NF-kB = nuclear factor kappa B; PSD95 = post-synaptic density protein 95; ONS = oxidative/nitrative stress.

Aquaporin-4 deletion in mice reduces encephalopathy and brain edema in experimental acute liver failure.

Brain edema and associated astrocyte swelling leading to increased intracranial pressure are hallmarks of acute liver failure (ALF). Elevated blood and brain levels of ammonia have been implicated in the development of brain edema in ALF. Cultured astrocytes treated with ammonia have been shown to undergo cell swelling and such swelling was associated with an increase in the plasma membrane expression of aquaporin-4 (AQP4) protein. Further, silencing the AQP4 gene in cultured astrocytes was shown to prevent the ammonia-induced cell swelling. Here, we examined the evolution of brain edema in AQP4-null mice and their wild type counterparts (WT-mice) in different models of ALF induced by thioacetamide (TAA) or acetaminophen (APAP). Induction of ALF with TAA or APAP significantly increased brain water content in WT mice (by 1.6% ± 0.3 and 2.3 ± 0.4%, respectively). AQP4 protein was significantly increased in brain plasma membranes of WT mice with ALF induced by either TAA or APAP. In contrast to WT-mice, brain water content did not increase in AQP4-null mice. Additionally, AQP4-null mice treated with either TAA or APAP showed a remarkably lesser degree of neurological deficits as compared to WT mice; the latter displayed an inability to maintain proper gait, and demonstrated a markedly reduced exploratory behavior, with the mice remaining in one corner of the cage with its head tilted downwards. These results support a central role of AQP4 in the brain edema associated with ALF.

Rac1b regulates NT3-stimulated Mek-Erk signaling, directing marrow-isolated adult multilineage inducible (MIAMI) cells toward an early neuronal phenotype.

Due to the limitations of neural stem cells to repair neuronal damage in the human brain, alternative approaches of repair using autologous adult stem cells have been examined for direct cell-replacement, or paracrine mediated neuroprotective effects. Human bone marrow-derived stromal cells (hMSCs) are a heterogeneous adult stem cell population with diverse immunomodulatory properties and the potential to differentiate into cells characteristic of all three germ layers. hMSCs are a renewable source of progenitor cells suitable for cell-based tissue repair. The marrow isolated adult multilineage inducible (MIAMI) cells developed by our laboratory are a developmentally immature homogeneous subpopulation of hMSCs that maintain self-renewal potential during ex vivo expansion, efficient differentiation capacity into neuron-like cells in vitro, as well as direct in vivo neuroprotection and functional recovery in animal models of neurological diseases. We now address the early signaling mechanisms regulating the neuron-like differentiation of MIAMI cells in vitro, in response to activation of the neurotrophic tyrosine-kinase receptor, type 3 (NTRK3) via neurotrophin 3 (NT3). We molecularly characterize a novel role for Rac1b mediating the neurogenic potential of MIAMI cells. Rac1b had an overall negative modulatory effect on the NT3-stimulated Mek1/2-Erk1/2 signaling pathway, proneuronal gene expression and neurite-like extensions. Rac1b was required for NT3-stimulated cell proliferation of MIAMI cells, yet was found to repress CCND1 and CCNB1 mRNA expression independent of NT3 stimulation, suggesting a dual neurotrophin dependent/independent function. Differential levels of Rac1b activity in hMSCs may explain the apparent contradictory reports regarding their neurogenic potential. These findings demonstrate the in vitro neurogenic potential of hMSCs as governed by Rac1b during NT3 stimulation.

Factors Influencing Telemedicine Use at a Northern New England Cancer Center During the COVID-19 Pandemic.

PURPOSE: To characterize the use of telemedicine for oncology care over the course of the COVID-19 pandemic in Northern New England with a focus on factors affecting trends. METHODS: We performed a retrospective observational study using patient visit data from electronic health records from hematology-oncology and radiation-oncology service lines spanning the local onset of the pandemic from March 18, 2020, through March 31, 2021. This period was subdivided into four phases designated as lockdown, transition, stabilization, and second wave. Generalized linear mixed regression models were used to estimate the effects of patient characteristics on trends for rates of telemedicine use across phases and the effects of visit type on patient satisfaction and postvisit ER or hospital admissions within 2 weeks. RESULTS: A total of 19,280 patients with 102,349 visits (13.1% audio-only and 1.4% video) were studied. Patient age (increased use in age < 45 and 85 years and older) and urban residence were associated with higher use of telemedicine, especially after initial lockdown. Recent cancer therapy, ER use, and hospital admissions in the past year were all associated with lower telemedicine utilization across pandemic phases. Provider clinical department corresponded to the largest differences in telemedicine use across all phases. ER and hospital admission rates in the 2 weeks after a telehealth visit were lower than those in in-person visits (0.7% v 1.3% and 1.2% v 2.7% for ER and hospital use, respectively; P < .001). Patient satisfaction did not vary across visit types. CONCLUSION: Telemedicine use in oncology during the COVID-19 pandemic varied according to the phase and patient, medical, and health system factors, suggesting opportunities for standardization of care and need for attention to equitable telemedicine access.

Adoption of Telemedicine in a Rural US Cancer Center Amid the COVID-19 Pandemic: Qualitative Study.

BACKGROUND: The COVID-19 pandemic necessitated a rapid shift to telemedicine to minimize patient and provider exposure risks. While telemedicine has been used in a variety of primary and specialty care settings for many years, it has been slow to be adopted in oncology care. Health care provider and administrator perspectives on factors affecting telemedicine use in oncology settings are not well understood, and the conditions associated with the COVID-19 pandemic offered the opportunity to study the adoption of telemedicine and the resulting provider and staff perspectives on its use. OBJECTIVE: The aim of this paper is to study the factors that influenced telemedicine uptake and sustained use in outpatient oncology clinics at a US cancer center to inform future telemedicine practices. METHODS: We used purposive sampling to recruit a mix of oncology specialty providers, practice managers, as well as nursing and administrative staff representing 5 outpatient oncology clinics affiliated with the Dartmouth Cancer Center, a large regional cancer center in the northeast of United States, to participate in semistructured interviews conducted over 6 weeks in spring 2021. The interview guide was informed by the 5 domains of the Consolidated Framework for Implementation Research, which include inner and outer setting factors, characteristics of the intervention (ie, telemedicine modality), individual-level factors (eg, provider and patient characteristics), and implementation processes. In total, 11 providers, 3 leaders, and 6 staff participated following verbal consent, and thematic saturation was reached across the full sample. We used a mixed deductive and inductive qualitative analysis approach to study the main influences on telemedicine uptake, implementation, and sustainability during the first year of the COVID-19 pandemic across the 5 settings. RESULTS: The predominant influencers of telemedicine adoption in this study were individual provider experiences and assumptions about patient preference and accessibility. Providers' early telemedicine experiences, especially if negative, influenced preferences for telephone over video and affected sustained use. Telemedicine was most favorably viewed for lower-acuity cancer care, visits less dependent on physical exam, and for patient and caregiver education. A lack of clinical champions, leadership guidance, and vision hindered the implementation of standardized practices and were cited as essential for telemedicine sustainability. Respondents expressed anxiety about sustaining telemedicine use if reimbursements for telephonic visits diminished or ceased. Opportunities to enhance future efforts include a need to provide additional guidance supporting telemedicine use cases and evidence of effectiveness in oncology care and to address provider concerns with communication quality. CONCLUSIONS: In a setting of decentralized care processes, early challenges in telemedicine implementation had an outsized impact on the nature and amount of sustained use. Proactively designed telemedicine care processes with attention to patient needs will be essential to support a sustained role for telemedicine in cancer care.

Na-K-Cl cotransporter-1 in the mechanism of cell swelling in cultured astrocytes after fluid percussion injury.

Brain edema and associated increased intracranial pressure are major consequences of traumatic brain injury (TBI). An important early component of the edema associated with TBI is astrocyte swelling (cytotoxic edema). Mechanisms for such swelling, however, are poorly understood. Ion channels/transporters/exchangers play a major role in cell volume regulation, and a disturbance in one or more of these systems may result in cell swelling. To examine potential mechanisms in TBI-mediated brain edema, we employed a fluid percussion model of in vitro barotrauma and examined the role of the ion transporter Na(+)-K(+)-2Cl(-)-cotransporter 1 (NKCC1) in trauma-induced astrocyte swelling as this transporter has been strongly implicated in the mechanism of cell swelling in various neurological conditions. Cultures exposed to trauma (3, 4, 5 atm pressure) caused a significant increase in NKCC1 activity (21%, 42%, 110%, respectively) at 3 h. At 5 atm pressure, trauma significantly increased NKCC1 activity at 1 h and it remained increased for up to 3 h. Trauma also increased the phosphorylation (activation) of NKCC1 at 1 and 3 h. Inhibition of MAPKs and oxidative/nitrosative stress diminished the trauma-induced NKCC1 phosphorylation as well as its activity. Bumetanide, an inhibitor of NKCC1, significantly reduced the trauma-induced astrocyte swelling (61%). Silencing NKCC1 with siRNA led to a reduction in trauma-induced NKCC1 activity as well as in cell swelling. These findings demonstrate the critical involvement of NKCC1 in the astrocyte swelling following in vitro trauma, and suggest that blocking NKCC1 activity may represent a useful therapeutic strategy for the cytotoxic brain edema associated with the early phase of TBI.

Neuroprotective properties of marrow-isolated adult multilineage-inducible cells in rat hippocampus following global cerebral ischemia are enhanced when complexed to biomimetic microcarriers.

Cell-based therapies for global cerebral ischemia represent promising approaches for neuronal damage prevention and tissue repair promotion. We examined the potential of marrow-isolated adult multilineage-inducible (MIAMI) cells, a homogeneous subpopulation of immature human mesenchymal stromal cell, injected into the hippocampus to prevent neuronal damage induced by global ischemia using rat organotypic hippocampal slices exposed to oxygen-glucose deprivation and rats subjected to asphyxial cardiac arrest. We next examined the value of combining fibronectin-coated biomimetic microcarriers (FN-BMMs) with epidermal growth factor (EGF)/basic fibroblast growth factor (bFGF) pre-treated MIAMI compared to EGF/bFGF pre-treated MIAMI cells alone, for their in vitro and in vivo neuroprotective capacity. Naïve and EGF/bFGF pre-treated MIAMI cells significantly protected the Cornu Ammonis layer 1 (CA1) against ischemic death in hippocampal slices and increased CA1 survival in rats. MIAMI cells therapeutic value was significantly increased when delivering the cells complexed with FN-BMMs, probably by increasing stem cell survival and paracrine secretion of pro-survival and/or anti-inflammatory molecules as concluded from survival, differentiation and gene expression analysis. Four days after oxygen and glucose deprivation and asphyxial cardiac arrest, few transplanted cells administered alone survived in the brain whereas stem cell survival improved when injected complexed with FN-BMMs. Interestingly, a large fraction of the transplanted cells administered alone or in complexes expressed ßIII-tubulin suggesting that partial neuronal transdifferentiation may be a contributing factor to the neuroprotective mechanism of MIAMI cells.

A multicenter comparison of established and emerging cardiac biomarkers for the diagnostic evaluation of chest pain in the emergency department.

BACKGROUND: The aim of this study is to assess the role of novel biomarkers for the diagnostic evaluation of acute coronary syndrome (ACS). METHODS: Among 318 patients presenting to an emergency department with acute chest discomfort, we evaluated the diagnostic value of 5 candidate biomarkers (amino terminal pro-B-type natriuretic peptide [NT-proBNP], ischemia modified albumin, heart fatty acid binding protein, high-sensitivity troponin I [hsTnI], and unbound free fatty acids [FFAu]) for detecting ACS, comparing their results with that of conventional troponin T (cTnT). RESULTS: Sixty-two subjects (19.5%) had ACS. The sensitivity and negative predictive values of NT-proBNP (73%, 90%) and hsTnI (57%, 89%) were higher than that of cTnT (22%, 84%). Unbound free fatty acids had the highest overall combination of sensitivity (75%), specificity (72%), and negative predictive values (92%) of all the markers examined. A significant increase in the C-statistic for cTnT resulted from the addition of results for NT-proBNP (change 0.09, P = .001), hsTnI (change 0.13, P < .001), and FFAu (change 0.15, P < .001). In integrated discrimination improvement and net reclassification improvement analyses, NT-proBNP, hsTnI, and FFAu added significant diagnostic information to cTnT; when changing the diagnostic criterion standard for ACS to hsTnI, FFAu still added significant reclassification for both events and nonevents. In serial sampling (n = 180), FFAu added important reclassification information to hsTnI. CONCLUSION: Among emergency department patients with symptoms suggestive of ACS, neither ischemia modified albumin nor heart fatty acid binding protein detected or excluded ACS, whereas NT-proBNP, hsTnI, or FFAu added diagnostic information to cTnT. In the context of hsTnI results, FFAu measurement significantly reclassified both false negatives and false positives at baseline and in serial samples.

EGF and bFGF pre-treatment enhances neural specification and the response to neuronal commitment of MIAMI cells.

AIMS: Multipotent mesenchymal stromal cells raise great interest for regenerative medicine studies. Some MSC subpopulations have the potential to undergo neural differentiation, including marrow isolated adult multilineage inducible (MIAMI) cells, which differentiate into neuron-like cells in a multi-step neurotrophin 3-dependent manner. Epidermal and basic fibroblast growth factors are often used in neuronal differentiation protocols for MSCs, but with a limited understanding of their role. In this study, we thoroughly assessed for the first time the capacity of these factors to enhance the neuronal differentiation of MSCs. MATERIALS AND METHODS: We have characterized MIAMI cell neuronal differentiation program in terms of stem cell molecule expression, cell cycle modifications, acquisition of a neuronal morphology and expression of neural and neuronal molecules in the absence and presence of an EGF-bFGF pre-treatment. RESULTS: EGF-bFGF pre-treatment down-regulated the expression of stemness markers Oct4A, Notch1 and Hes5, whereas neural/neuronal molecules Nestin, Pax6, Ngn2 and the neurotrophin receptor tyrosine kinase 1 and 3 were up-regulated. During differentiation, a sustained Erk phosphorylation in response to NT3 was observed, cells began to exit from the cell cycle and exhibit increased neurite-like extensions. In addition, neuronal ß3-tubulin and neurofilament expression was increased; an effect mediated via the Erk pathway. A slight pre-oligodendrocyte engagement was noted, and no default neurotransmitter phenotype was observed. Overall, mesodermal markers were unaffected or decreased, while neurogenic/adipogenic PPARγ2 was increased. CONCLUSION: EGF and bFGF pre-treatment enhances neural specification and the response to neuronal commitment of MIAMI cells, further increasing their potential use in adult cell therapy of the nervous system.

Aquaporin-4 in manganese-treated cultured astrocytes.

Manganese in excess is neurotoxic and causes CNS injury resembling that of Parkinson's disease. In brain, astrocytes predominantly take up and accumulate manganese and are thus vulnerable to its toxicity. Manganese was shown to induce cell swelling in cultured astrocytes, and oxidative/nitrosative stress (ONS) mediates such swelling. As aquaporin-4 (AQP4) is important in the mechanism of astrocyte swelling, we examined the effect of manganese on AQP4 protein levels in cultured astrocytes. Treatment of cultures with manganese increased AQP4 protein in the plasma membrane (PM), whereas total cellular AQP4 protein and mRNA levels were unchanged, suggesting that increased AQP4 levels is due to its increased stability and/or increased trafficking to the PM and not to its neosynthesis. AQP4 gene silencing by small interfering ribonucleic acid resulted in a marked reduction in astrocyte swelling by manganese. Antioxidants, as well as an inhibitor of nitric oxide synthase, diminished the increase in AQP4 protein expression, suggesting a role of ONS in the mechanism of AQP4 increase. As ONS is known to activate mitogen-activated protein kinases (MAPKs) and MAPK activation has been implicated in astrocyte swelling, we examined the effect of manganese on the activation of MAPKs and found an increased phosphorylation of extracellular signal-regulated kinase (ERK)1/2, C-Jun amino-terminal kinase (JNK)1/2/3, and p38-MAPK. Inhibitors of ERK1/2 and p38-MAPK (but not of JNK) blocked (40-60%) the manganese-induced increase in AQP4 protein content and astrocyte swelling, suggesting the involvement of these kinases in the increased AQP4 content. Inhibition of oxidative stress or MAPKs may represent potential strategies for counteracting AQP4-related neurological complications associated with manganese toxicity.

EF1alpha and RPL13a represent normalization genes suitable for RT-qPCR analysis of bone marrow derived mesenchymal stem cells.

BACKGROUND: RT-qPCR analysis is a widely used method for the analysis of mRNA expression throughout the field of mesenchymal stromal cell (MSC) research. Comparison between MSC studies, both in vitro and in vivo, are challenging due to the varied methods of RT-qPCR data normalization and analysis. Therefore, this study focuses on putative housekeeping genes for the normalization of RT-qPCR data between heterogeneous commercially available human MSC, compared with more homogeneous populations of MSC such as MIAMI and RS-1 cells. RESULTS: Eight genes including; ACTB, B2M, EF1alpha, GAPDH, RPL13a, YWHAZ, UBC and HPRT1 were tested as possible housekeeping genes based on their expression level and variability. EF1alpha and RPL13a were validated for RT-qPCR analysis of MIAMI cells during expansion in varied oxygen tensions, endothelial differentiation, neural precursor enrichment, and during the comparison with RS-1 cells and commercially available MSC. RPL13a and YWHAZ were validated as normalization genes for the cross-species analysis of MIAMI cells in an animal model of focal ischemia. GAPDH, which is one of the most common housekeeping genes used for the normalization of RT-qPCR data in the field of MSC research, was found to have the highest variability and deemed not suitable for normalization of RT-qPCR data. CONCLUSIONS: In order to make comparisons between heterogeneous MSC populations, as well as adult stem cell like MSC which are used in different laboratories throughout the world, it is important to have a standardized, reproducible set of housekeeping genes for RT-qPCR analysis. In this study we demonstrate that EF1alpha, RPL13a and YWHAZ are suitable genes for the RT-qPCR analysis and comparison of several sources of human MSC during in vitro characterization and differentiation as well as in an ex vivo animal model of global cerebral ischemia. This will allow for the comparative RT-qPCR analysis of multiple MSC populations with the goal of future use in animal models of disease as well as tissue repair.

A fentanyl-based pain management protocol provides early analgesia for adult trauma patients.

BACKGROUND: In the past two decades, a number of reports have identified inadequate treatment of pain among emergency department patients. No study has evaluated the frequency or effectiveness of early analgesia in the trauma patient. The objective of this study was to determine the effect of a protocol-driven pain management scheme on time to initiation of analgesia among trauma patients. METHODS: A fentanyl-based protocol was developed with patients being assigned to one of three treatment arms based on hemodynamics and Glasgow Coma Scale (GCS) score. Using an institutional review board-approved before and after study design, patients over the age of 14 and meeting trauma system activation criteria at the Dartmouth-Hitchcock Medical Center were eligible. Results were compared with a retrospective chart review of eligible patients treated during a matched preprotocol time period in 2002. The primary outcome measure was time to initiation of analgesia. Secondary outcome measures included (1) the proportion of patients receiving their first analgesia dose within 30 minutes, (2) the number of patients receiving multiple doses of analgesia in the trauma bay, and (3) adverse events. Pain level was assessed using either a Numeric Pain Scale (for patients with a GCS score of 15) or a Behavioral Pain Assessment Scale (GCS score <15). RESULTS: Implementation of the protocol resulted in a decrease in the mean time to initiation of analgesia from 53.61 minutes +/- 6.88 minutes to 27.94 minutes +/- 3.34 minutes (p = 0.001). The protocol also increased the percentage of patients receiving analgesia within the first 30 minutes of arrival from 44.4% to 74.6% (p < 0.001). There were no differences between the two groups in terms of baseline characteristics or adverse events. CONCLUSIONS: The implementation of a fentanyl-based pain management protocol resulted in a marked reduction in time to initial analgesia among trauma patients. There was no evidence of an increase in adverse events. This tool has the potential to be easily extrapolated and applied to other trauma systems.

The lack of an adult seat belt law in New Hampshire: live free and die?

New Hampshire (NH) is the only state with no adult seat belt law. The lack of a restraint law may stem from a reluctance to infringe on individual freedoms. Dartmouth-Hitchcock Medical Center (DHMC) is located in NH, only 4 miles from the Vermont (VT) border. As a result, residents of both states are well represented in the Emergency Department (ED) population. This provided an opportunity to investigate the relationship between opinions, behaviors and the presence or absence of a restraint law. Adult DHMC ED patients were surveyed with respect to seat belt use in states with and without restraint laws, risk taking behaviors, and feelings of infringement on personal freedom. The results demonstrated no difference in restraint use between NH and VT residents while traveling in a state with a seat belt law. However, significantly fewer NH residents reported restraint use when traveling in a state without a seat belt law. These dissimilarities were not explained by differences in risk taking behaviors or by differences in feelings of infringement on freedom. This suggests that actual seat belt use reflects adherence to the law rather than concerns over personal freedom. This may inspire a reassessment of the acceptability of an adult restraint law in New Hampshire.

Hepatocyte growth factor and alternative splice variants - expression, regulation and implications in osteogenesis and bone health and repair.

INTRODUCTION: Bone marrow-derived mesenchymal stem cells (MSCs) can differentiate into multiple cell types, including osteoblasts, chondrocytes, and adipocytes. These pluripotent cells secrete hepatocyte growth factor (HGF), which regulates cell growth, survival, motility, migration, mitogenesis and is important for tissue development/regeneration. HGF has four splice variants, NK1, NK2, NK3, and NK4 which have varying functions and affinities for the HGF receptor, cMET. HGF promotes osteoblastic differentiation of MSCs into bone forming cells, playing a role in bone development, health and repair. AREAS COVERED: This review will focus on the effects of HGF in osteogenesis, bone repair and bone health, including structural and functional insights into the role of HGF in the body. EXPERT OPINION: Approximately 6.2 million Americans experience a fracture annually, with 5-10% being mal- or non-union fractures. HGF is important in priming MSCs for osteogenic differentiation in vitro and is currently being studied to assess its role during bone repair in vivo. Due to the high turnover rate of systemic HGF, non-classic modes of HGF-treatment, including naked-plasmid HGF delivery and the use of HGF splice variants (NK1 & NK2) are being studied to find safe and efficacious treatments for bone disorders, such as mal- or non-union fractures.

Decreased STAT3 Phosphorylation Mediates Cell Swelling in Ammonia-Treated Astrocyte Cultures.

Brain edema, due largely to astrocyte swelling, and the subsequent increase in intracranial pressure and brain herniation, are major complications of acute liver failure (ALF). Elevated level of brain ammonia has been strongly implicated in the development of astrocyte swelling associated with ALF. The means by which ammonia brings about astrocyte swelling, however, is incompletely understood. Recently, oxidative/nitrosative stress and associated signaling events, including activation of mitogen-activated protein kinases (MAPKs), as well as activation of the transcription factor, nuclear factor-kappaB (NF-κB), have been implicated in the mechanism of ammonia-induced astrocyte swelling. Since these signaling events are known to be regulated by the transcription factor, signal transducer and activator of transcription 3 (STAT3), we examined the state of STAT3 activation in ammonia-treated cultured astrocytes, and determined whether altered STAT3 activation and/or protein expression contribute to the ammonia-induced astrocyte swelling. STAT3 was found to be dephosphorylated (inactivated) at Tyrosine705 in ammonia-treated cultured astrocytes. Total STAT3 protein level was also reduced in ammonia-treated astrocytes. We also found a significant increase in protein tyrosine phosphatase receptor type-1 (PTPRT-1) protein expression in ammonia-treated cultured astrocytes, and that inhibition of PTPRT-1 enhanced the phosphorylation of STAT3 after ammonia treatment. Additionally, exposure of cultured astrocytes to inhibitors of protein tyrosine phosphatases diminished the ammonia-induced cell swelling, while cultured astrocytes over-expressing STAT3 showed a reduction in the astrocyte swelling induced by ammonia. Collectively, these studies strongly suggest that inactivation of STAT3 represents a critical event in the mechanism of the astrocyte swelling associated with acute liver failure.