Long-term reversal of chronic pain behavior in rodents through elevation of spinal agmatine.

Chronic pain remains a significant burden worldwide, and treatments are often limited by safety or efficacy. The decarboxylated form of L-arginine, agmatine, antagonizes N-methyl-d-aspartate receptors, inhibits nitric oxide synthase, and reverses behavioral neuroplasticity. We hypothesized that expressing the proposed synthetic enzyme for agmatine in the sensory pathway could reduce chronic pain without motor deficits. Intrathecal delivery of an adeno-associated viral (AAV) vector carrying the gene for arginine decarboxylase (ADC) prevented the development of chronic neuropathic pain as induced by spared nerve injury in mice and rats and persistently reversed established hypersensitivity 266 days post-injury. Spinal long-term potentiation was inhibited by both exogenous agmatine and AAV-human ADC (hADC) vector pre-treatment but was enhanced in rats treated with anti-agmatine immunoneutralizing antibodies. These data suggest that endogenous agmatine modulates the neuroplasticity associated with chronic pain. Development of approaches to access this inhibitory control of neuroplasticity associated with chronic pain may yield important non-opioid pain-relieving options.

Lower SARS-CoV-2-specific humoral immunity in people living with HIV-1 recovered from nonhospitalized COVID-19.

People living with HIV-1 (PLWH) exhibit more rapid antibody decline following routine immunization and elevated baseline chronic inflammation than people without HIV-1 (PWOH), indicating potential for diminished humoral immunity during SARS-CoV-2 infection. Conflicting reports have emerged on the ability of PLWH to maintain humoral protection against SARS-CoV-2 coinfection during convalescence. It is unknown whether peak COVID-19 severity, along with HIV-1 infection status, associates with the quality and quantity of humoral immunity following recovery. Using a cross-sectional observational cohort from the United States and Peru, adults were enrolled 1-10 weeks after SARS-CoV-2 infection diagnosis or symptom resolution. Serum antibodies were analyzed for SARS-CoV-2-specific response rates, binding magnitudes, ACE2 receptor blocking, and antibody-dependent cellular phagocytosis. Overall, (a) PLWH exhibited a trend toward decreased magnitude of SARS-CoV-2-specific antibodies, despite modestly increased overall response rates when compared with PWOH; (b) PLWH recovered from symptomatic outpatient COVID-19 had comparatively diminished immune responses; and (c) PLWH lacked a corresponding increase in SARS-CoV-2 antibodies with increased COVID-19 severity when asymptomatic versus symptomatic outpatient disease was compared.

Impact of Interhospital Transfer on Outcomes in Non-emergency Colorectal Surgery.

BACKGROUND: A paucity of data exists on the impact of transfer status on outcomes for patients undergoing non-emergency (urgent) colorectal surgery. This study characterized transferred patients undergoing urgent colorectal surgery and determined which patient comorbidities significantly contributed to poor outcomes. METHODS: The American College of Surgeons National Surgical Quality Improvement Program database from 2012 to 2013 was used. Urgent direct admissions undergoing colon, rectum, or small bowel operations were compared to urgent transfers using bivariate and multivariable analysis models. Primary outcomes were overall complications, hospital length of stay, and mortality. RESULTS: A total of 82,151 admissions were analyzed. After multivariable analysis, direct admission patients had nearly similar risk of complications (RR = 0.95; 95% CI 0.91-0.99) and length of hospital stay (7% shorter; 95% CI 4-9%), as well as no difference in mortality (RR = 0.94; 95% CI 0.80-1.11). CONCLUSIONS: Transfer status alone confers minimal risk toward higher complication rates and longer hospital length of stay in patients undergoing urgent colorectal surgery, and the poor outcomes observed in this cohort are largely due to patient comorbidities and disease severity. Our results suggest that outcomes in transferred colorectal surgery patients undergoing urgent operations depend mainly on operative acuity and clinical factors, and to a lesser degree transfer status.

Supraspinal gene transfer by intrathecal adeno-associated virus serotype 5.

We report the pattern of transgene expression across brain regions after intrathecal delivery of adeno-associated virus serotype 5 (AAV5). Labeling in hindbrain appeared to be primarily neuronal, and was detected in sensory nuclei of medulla, pontine nuclei, and all layers of cerebellar cortex. Expression in midbrain was minimal, and generally limited to isolated neurons and astrocytes in the cerebral peduncles. GFP immunoreactivity (-ir) in thalamus was most prominent in medial geniculate nucleus, and otherwise limited to posterior nuclei of the dorsal and lateral margins. Labeling was also observed in neurons and astrocytes of the hippocampal formation and amygdaloid complex. In the hippocampal formation, GFP-ir was found in neuronal cell bodies of the rostral ventral portion, but was largely restricted to fiber-like staining in the molecular layer of dentate gyrus and stratum lacunosum-moleculare of the rostral dorsal region. GFP-ir was seen in neurons and astroglia throughout caudal cortex, whereas in rostral regions of neocortex it was limited to isolated neurons and non-neuronal cells. Labeling was also present in olfactory bulb. These results demonstrate that intrathecal delivery of AAV5 vector leads to transgene expression in discrete CNS regions throughout the rostro-caudal extent of the neuraxis. A caudal-to-rostral gradient of decreasing GFP-ir was present in choroid plexus and Purkinje cells, suggesting that spread of virus through cerebrospinal fluid plays a role in the resulting transduction pattern. Other factors contributing to the observed expression pattern likely include variations in cell-surface receptors and inter-parenchymal space.

Biodistribution of adeno-associated virus serotype 9 (AAV9) vector after intrathecal and intravenous delivery in mouse.

Adeno-associated virus serotype 9 (AAV9)-mediated gene transfer has been reported in central nervous system (CNS) and peripheral tissues. The current study compared the pattern of expression of Green Fluorescent Protein (GFP) across the mouse CNS and selected peripheral tissues after intrathecal (i.t.) or intravenous (i.v.) delivery of equivalent doses of single-stranded AAV9 vector. After i.t. delivery, GFP immunoreactivity (-ir) was observed in spinal neurons, primary afferent fibers and corresponding primary sensory neurons at all spinal levels. Robust transduction was seen in small and large dorsal root ganglion (DRG) neurons as well as trigeminal and vagal primary afferent neurons. Transduction efficiency in sensory ganglia was substantially lower in i.v. treated mice. In brain, i.v. delivery yielded GFP-immunoreactivity (-ir) primarily in spinal trigeminal tract, pituitary, and scattered isolated neurons and astrocytes. In contrast, after i.t. delivery, GFP-ir was widespread throughout CNS, with greater intensity and more abundant neuropil-like staining at 6 weeks compared to 3 weeks. Brain regions with prominent GFP-ir included cranial nerve nuclei, ventral pons, cerebellar cortex, hippocampus, pituitary, choroid plexus, and selected nuclei of midbrain, thalamus and hypothalamus. In cortex, GFP-ir was associated with blood vessels, and was seen in both neurons and astrocytes. In the periphery, GFP-ir in colon and ileum was present in the enteric nervous system in both i.v. and i.t. treated mice. Liver and adrenal cortex, but not adrenal medulla, also showed abundant GFP-ir after both routes of delivery. In summary, i.t. delivery yielded higher transduction efficiency in sensory neurons and the CNS. The observation of comparable gene transfer to peripheral tissues using the two routes indicates that a component of i.t. delivered vector is redistributed from the subarachnoid space to the systemic circulation.

Protein kinase Cε is required for spinal analgesic synergy between delta opioid and alpha-2A adrenergic receptor agonist pairs.

We recently showed that spinal synergistic interactions between δ opioid receptors (δORs) and α2A adrenergic receptors (α2AARs) require protein kinase C (PKC). To identify which PKC isoforms contribute to analgesic synergy, we evaluated the effects of various PKC-isoform-specific peptide inhibitors on synergy between δORs and α2AARs using the tail flick assay of thermal nociception in mice. Only a PKCε inhibitor abolished synergy between a δOR agonist and an α2AAR agonist. We tested a panel of combinations of opioid and adrenergic agonists in PKCε knock-out mice and found that all four combinations of a δOR agonist and an α2AAR agonist required PKCε for antinociceptive synergy. None of the combinations of a µOR agonist with an α2AR agonist required PKCε. Immunohistochemistry confirmed that PKCε could be found in the population of peptidergic primary afferent nociceptors where δORs and α2AARs have been found to extensively colocalize. Immunoreactivity for PKCε was found in the majority of dorsal root ganglion neurons and intensely labeled laminae I and II of the spinal cord dorsal horn. PKCε is widespread in the spinal nociceptive system and in peptidergic primary afferents it appears to be specifically involved in mediating the synergistic interaction between δORs and α2AARs.

Cardioprotective mechanisms activated in response to myocardial ischemia.

Myocardial ischemia, a disorder causing myocardial infarction and malfunction, can activate various adaptive mechanisms that protect cardiomyocytes from ischemic injury. During the early hours post myocardial ischemia, injured cardiac cells can release several molecules, including adenosine, opioids, and bradykinin, which promote myocardial survival by activating the G protein signaling pathways. During a later phase about several days, myocardial ischemia induces upregulation of growth factors and cytokines, including VEGF, ILGF, HGF, and SDF-1, in the injured myocardium, contributing to cardioprotection. In addition to the injured heart, the liver participates in cardioprotection. In response to myocardial ischemia, the liver upregulates and releases secretory proteins, including FGF21 and TFF3, both of which promote cardiomyocyte survival. The liver also provides a reservoir of hepatic cells that mobilize to the site of myocardial ischemia, potentially contributing to cardioprotection. Taken together, the early and late mechanisms act coordinately in a time-dependent manner, ensuring effective cardioprotection post myocardial infarction. Investigations on these innate cardioprotective mechanisms have provided insights into the development of cardioprotective strategies for treating myocardial infarction. In this article, the authors review the innate mechanisms of cardioprotection in myocardial ischemia.

Differential adeno-associated virus mediated gene transfer to sensory neurons following intrathecal delivery by direct lumbar puncture.

BACKGROUND: Neuronal transduction by adeno-associated viral (AAV) vectors has been demonstrated in cortex, brainstem, cerebellum, and sensory ganglia. Intrathecal delivery of AAV serotypes that transduce neurons in dorsal root ganglia (DRG) and spinal cord offers substantial opportunities to 1) further study mechanisms underlying chronic pain, and 2) develop novel gene-based therapies for the treatment and management of chronic pain using a non-invasive delivery route with established safety margins. In this study we have compared expression patterns of AAV serotype 5 (AAV5)- and AAV serotype 8 (AAV8)-mediated gene transfer to sensory neurons following intrathecal delivery by direct lumbar puncture. RESULTS: Intravenous mannitol pre-treatment significantly enhanced transduction of primary sensory neurons after direct lumbar puncture injection of AAV5 (rAAV5-GFP) or AAV8 (rAAV8-GFP) carrying the green fluorescent protein (GFP) gene. The presence of GFP in DRG neurons was consistent with the following evidence for primary afferent origin of the majority of GFP-positive fibers in spinal cord: 1) GFP-positive axons were evident in both dorsal roots and dorsal columns; and 2) dorsal rhizotomy, which severs the primary afferent input to spinal cord, abolished the majority of GFP labeling in dorsal horn. We found that both rAAV5-GFP and rAAV8-GFP appear to preferentially target large-diameter DRG neurons, while excluding the isolectin-B4 (IB4) -binding population of small diameter neurons. In addition, a larger proportion of CGRP-positive cells was transduced by rAAV5-GFP, compared to rAAV8-GFP. CONCLUSIONS: The present study demonstrates the feasibility of minimally invasive gene transfer to sensory neurons using direct lumbar puncture and provides evidence for differential targeting of subtypes of DRG neurons by AAV vectors.

Supraspinally-administered agmatine attenuates the development of oral fentanyl self-administration.

The decarboxylation product of arginine, agmatine, has effectively reduced or prevented opioid-induced tolerance and dependence when given either systemically (intraperitoneally or subcutaneously) or centrally (intrathecally or intracerebroventricularly). Systemically administered agmatine also reduces the escalation phase of intravenous fentanyl self-administration in rats. The present study assessed whether centrally (intracerebroventricular, i.c.v.) delivered agmatine could prevent the development of fentanyl self-administration in mice. Mice were trained to respond under a fixed-ratio 1 (FR1) schedule for either fentanyl (0.7 microg/70 microl, p.o.) or food reinforcement. Agmatine (10 nmol/5 microl), injected i.c.v. 12-14 h before the first session and every other evening (12-14 h before session) for 2 weeks, completely attenuated oral fentanyl self-administration (but not food-maintained responding) compared to saline-injected controls. When agmatine was administered after fentanyl self-administration had been established (day 8) it had no attenuating effects on bar pressing. This dose of agmatine does not decrease locomotor activity as assessed by rotarod. The present findings significantly extend the previous observation that agmatine prevents opioid-maintained behavior to a chronic model of oral fentanyl self-administration as well as identifying a supraspinal site of action for agmatine inhibition of drug addiction.