ß-endorphin at the intersection of pain and cancer progression: Preclinical evidence.

We examined the association between endogenous opioid ß-endorphin, cancer progression and pain in a transgenic mouse model of breast cancer, with a rat C3(1) simian virus 40 large tumor antigen fusion gene (C3TAg). C3TAg mice develop ductal epithelial atypia at 8 weeks, progression to intra-epithelial neoplasia at 12 weeks, and invasive carcinoma with palpable tumors at 16 weeks. Consistent with invasive carcinoma at 4 months of age, C3TAg mice demonstrate a significant increase in hyperalgesia compared to younger C3TAg or control FVBN mice without tumors. Our data show that the growing tumor contributes to circulating ß-endorphin. As an endogenous ligand of mu opioid receptor, ß-endorphin has analgesic activity. Paradoxically, we observed an increase in pain in transgenic breast cancer mice with significantly high circulating and tumor-associated ß-endorphin. Increased circulating ß-endorphin correlates with increasing tumor burden. ß-endorphin induced the activation of mitogenic and survival-promoting signaling pathways, MAPK/ERK 1/2, STAT3 and Akt, observed by us in human MDA-MB-231 cells suggesting a role for ß-endorphin in breast cancer progression and associated pain.

Considerations for Cannabis Use to Treat Pain in Sickle Cell Disease.

Pain in Sickle Cell Disease (SCD) is a major comorbidity and unique with acute pain due to recurrent and episodic vaso-occlusive crises as well as chronic pain, which can span an individual's entire life. Opioids are the mainstay treatment for pain in SCD. Due to recent health crises raised by adverse effects including deaths from opioid use, pain management in SCD is adversely affected. Cannabis and its products are most widely used for pain in multiple conditions and also by patients with SCD on their own. With the availability of "Medical Cannabis" and approval to use cannabis as medicine across majority of States in the United States as well as over-the-counter preparations, cannabis products are being used increasingly for SCD. The reliability of many of these products remains questionable, which poses a major health risk to the vulnerable individuals seeking pain relief. Therefore, this review provides up to date insights into available categories of cannabis-based treatment strategies, their mechanism of action and pre-clinical and clinical outcomes in SCD. It provides evidence for the benefits and risks of cannabis use in SCD and cautions about the unreliable and unvalidated products that may be adulterated with life-threatening non-cannabis compounds.

Spatiotemporal Alterations in Gait in Humanized Transgenic Sickle Mice.

Sickle cell disease (SCD) is a hemoglobinopathy affecting multiple organs and featuring acute and chronic pain. Purkinje cell damage and hyperalgesia have been demonstrated in transgenic sickle mice. Purkinje cells are associated with movement and neural function which may influence pain. We hypothesized that Purkinje cell damage and/or chronic pain burden provoke compensatory gait changes in sickle mice. We found that Purkinje cells undergoe increased apoptosis as shown by caspase-3 activation. Using an automated gait measurement system, MouseWalker, we characterized spatiotemporal gait characteristics of humanized transgenic BERK sickle mice in comparison to control mice. Sickle mice showed alteration in stance instability and dynamic gait parameters (walking speed, stance duration, swing duration and specific swing indices). Differences in stance instability may reflect motor dysfunction due to damaged Purkinje cells. Alterations in diagonal and all stance indices indicative of hesitation during walking may originate from motor dysfunction and/or arise from fear and/or anticipation of movement-evoked pain. We also demonstrate that stance duration, diagonal swing indices and all stance indices correlate with both mechanical and deep tissue hyperalgesia, while stance instability correlates with only deep tissue hyperalgesia. Therefore, objective analysis of gait in SCD may provide insights into neurological impairment and pain states.

Self-Assembled 3D Nanosplit Rings for Plasmon-Enhanced Optofluidic Sensing.

Plasmonic sensors are commonly defined on two-dimensional (2D) surfaces with an enhanced electromagnetic field only near the surface, which requires precise positioning of the targeted molecules within hotspots. To address this challenge, we realize segmented nanocylinders that incorporate plasmonic (1-50 nm) gaps within three-dimensional (3D) nanostructures (nanocylinders) using electron irradiation triggered self-assembly. The 3D structures allow desired plasmonic patterns on their inner cylindrical walls forming the nanofluidic channels. The nanocylinders bridge nanoplasmonics and nanofluidics by achieving electromagnetic field enhancement and fluid confinement simultaneously. This hybrid system enables rapid diffusion of targeted species to the larger spatial hotspots in the 3D plasmonic structures, leading to enhanced interactions that contribute to a higher sensitivity. This concept has been demonstrated by characterizing an optical response of the 3D plasmonic nanostructures using surface-enhanced Raman spectroscopy (SERS), which shows enhancement over a 22 times higher intensity for hemoglobin fingerprints with nanocylinders compared to 2D nanostructures.

Microfluidics in Sickle Cell Disease Research: State of the Art and a Perspective Beyond the Flow Problem.

Sickle cell disease (SCD) is the monogenic hemoglobinopathy where mutated sickle hemoglobin molecules polymerize to form long fibers under deoxygenated state and deform red blood cells (RBCs) into predominantly sickle form. Sickled RBCs stick to the vascular bed and obstruct blood flow in extreme conditions, leading to acute painful vaso-occlusion crises (VOCs) - the leading cause of mortality in SCD. Being a blood disorder of deformed RBCs, SCD manifests a wide-range of organ-specific clinical complications of life (in addition to chronic pain) such as stroke, acute chest syndrome (ACS) and pulmonary hypertension in the lung, nephropathy, auto-splenectomy, and splenomegaly, hand-foot syndrome, leg ulcer, stress erythropoiesis, osteonecrosis and osteoporosis. The physiological inception for VOC was initially thought to be only a fluid flow problem in microvascular space originated from increased viscosity due to aggregates of sickled RBCs; however, over the last three decades, multiple molecular and cellular mechanisms have been identified that aid the VOC in vivo. Activation of adhesion molecules in vascular endothelium and on RBC membranes, activated neutrophils and platelets, increased viscosity of the blood, and fluid physics driving sickled and deformed RBCs to the vascular wall (known as margination of flow) - all of these come together to orchestrate VOC. Microfluidic technology in sickle research was primarily adopted to benefit from mimicking the microvascular network to observe RBC flow under low oxygen conditions as models of VOC. However, over the last decade, microfluidics has evolved as a valuable tool to extract biophysical characteristics of sickle red cells, measure deformability of sickle red cells under simulated oxygen gradient and shear, drug testing, in vitro models of intercellular interaction on endothelialized or adhesion molecule-functionalized channels to understand adhesion in sickle microenvironment, characterizing biomechanics and microrheology, biomarker identification, and last but not least, for developing point-of-care diagnostic technologies for low resource setting. Several of these platforms have already demonstrated true potential to be translated from bench to bedside. Emerging microfluidics-based technologies for studying heterotypic cell-cell interactions, organ-on-chip application and drug dosage screening can be employed to sickle research field due to their wide-ranging advantages.

Pain and sickle cell disease.

PURPOSE OF REVIEW: Pain is a major comorbidity of sickle cell disease (SCD). Opioids are the mainstay for pain treatment but remain suboptimal. We discuss mechanism-based treatable targets devoid of opioids to prevent and/or treat SCD pain. RECENT FINDINGS: Understanding the pathogenesis of pain is critical to develop targeted therapies. Nevertheless, acute and chronic pain can have independent and/or overlapping mechanisms. The origin of pain involves neurovascular and neuroimmune interactions from the periphery and/or central nervous system. Immunomodulatory components of acute and/or chronic sickle pain for targeting/preventing pain genesis include mast cell and microglial activation, neurogenic inflammation, and leukocyte-derived elastase. Vascular modulators include hypoxia/reperfusion injury, oxidative stress, hemolysis, and adhesion molecules. However, existent pain requires analgesics devoid of an inadvertent effect on sickle pathobiology. Recent analgesic targets include cannabinoid and nociceptin receptors and serotonergic spinothalamic pathway. Complementary approaches (e.g., acupuncture, hypnosis, perception-based therapies) have shown analgesic potential. Owing to heterogeneity in pain development, it remains challenging to combat SCD pain with any one therapy. SUMMARY: SCD pain involves neuroimmune and neurovascular interactions. Such interactions have pronociceptive impacts and impart therapy resistance. Elucidating molecular and cellular entities affecting neuronal interactions in sickle microenvironment may prevent SCD pain and/or provide improved analgesic approaches.

Transcriptomic analysis of gene signatures associated with sickle pain.

Pain is a hallmark feature of sickle cell disease (SCD). Recurrent and unpredictable acute pain due to vaso-oclussive crises (VOC) is unique to SCD; and can be superimposed on chronic pain. To examine the mechanisms underlying pain in SCD, we performed RNA sequencing of dorsal root ganglion (DRG) of transgenic sickle mice and their age-matched control mice expressing normal human hemoglobin A, at 2 and 5 months of age. Sickle and control mice of both ages were equally divided into hypoxia/reoxygenation (to simulate VOC) and normoxia treatment, resulting in eight groups of mice. Each group had at least six mice. RNA isolated from the DRG was sequenced and paired-end 50 bp sequencing data were generated using Illumina's HiSeq 2000. This large dataset can serve as a resource for examining transcriptional changes in the DRG that are associated with age and hypoxia/reoxygenation associated signatures of nociceptive mechanisms underlying chronic and acute pain, respectively.

Mast Cell-Mediated Mechanisms of Nociception.

Mast cells are tissue-resident immune cells that release immuno-modulators, chemo-attractants, vasoactive compounds, neuropeptides and growth factors in response to allergens and pathogens constituting a first line of host defense. The neuroimmune interface of immune cells modulating synaptic responses has been of increasing interest, and mast cells have been proposed as key players in orchestrating inflammation-associated pain pathobiology due to their proximity to both vasculature and nerve fibers. Molecular underpinnings of mast cell-mediated pain can be disease-specific. Understanding such mechanisms is critical for developing disease-specific targeted therapeutics to improve analgesic outcomes. We review molecular mechanisms that may contribute to nociception in a disease-specific manner.

The free heme concentration in healthy human erythrocytes.

Heme, the prosthetic group of hemoglobin, may be released from its host due to an intrinsic instability of hemoglobin and accumulate in the erythrocytes. Free heme is in the form of hematin (Fe(3+) protoporphyrin IX OH) and follows several pathways of biochemical toxicity to tissues, cells, and organelles since it catalyzes the production of reactive oxygen species. To determine concentration of soluble free heme in human erythrocytes, we develop a new method. We lyse the red blood cells and isolate free heme from hemoglobin by dialysis. We use the heme to reconstitute horseradish peroxidase (HRP) from an excess of the apoenzyme and determine the HRP reaction rate from the evolution of the emitted luminescence. We find that in a population of five healthy adults the average free heme concentration in the erythrocytes is 21±2µM, ca. 100× higher than previously determined. Tests suggest that the lower previous value was due to the use of elevated concentrations of NaCl, which drive hematin precipitation and re-association with apoglobin. We show that the found hematin concentration is significantly higher than estimates based on equilibrium release and the known hematin dimerization. The factors that lead to enhanced heme release remain an open question.