Effect of stretching on inflammation in a subcutaneous carrageenan mouse model analyzed at single-cell resolution.

Understanding the factors that influence the biological response to inflammation is crucial, due to its involvement in physiological and pathological processes, including tissue repair/healing, cancer, infections, and autoimmune diseases. We have previously demonstrated that in vivo stretching can reduce inflammation and increase local pro-resolving lipid mediators in rats, suggesting a direct mechanical effect on inflammation resolution. Here we aimed to explore further the effects of stretching at the cellular/molecular level in a mouse subcutaneous carrageenan-inflammation model. Stretching for 10 min twice a day reduced inflammation, increased the production of pro-resolving mediator pathway intermediate 17-HDHA at 48 h postcarrageenan injection, and decreased both pro-resolving and pro-inflammatory mediators (e.g., PGE2 and PGD2 ) at 96 h. Single-cell RNA sequencing analysis of inflammatory lesions at 96 h showed that stretching increased the expression of both pro-inflammatory (Nos2) and pro-resolution (Arg1) genes in M1 and M2 macrophages at 96 h. An intercellular communication analysis predicted specific ligand-receptor interactions orchestrated by neutrophils and M2a macrophages, suggesting a continuous neutrophil presence recruiting immune cells such as activated macrophages to contain the antigen while promoting resolution and preserving tissue homeostasis.

Flexibility and strength training in asthma: A pilot study.

OBJECTIVE: Although less is known about musculoskeletal factors that may contribute to asthma symptoms, body-based treatments addressing movement restrictions of the chest and shoulders may be a useful adjunct to asthma pharmacotherapy. In this pilot study, we compared asthma symptoms, pulmonary function tests, and medication use before and after a course of resistance flexibility and strength training (RFST) treatments in human subjects with asthma. METHODS: Patients with asthma (n = 10; mean age 23 years) completed questionnaires (Asthma Control Questionnaire (ACQ), Asthma Quality of Life Questionnaire (AQLQ), Asthma Control Test (ACT)), spirometry, chest wall excursion, and shoulder range of motion (ROM) before and after a series of four RFST treatments over 47 ± 21 days. Each treatment consisted of a one-hour session involving eccentric stretching of the arm, shoulder, and chest while lying in a supine position. RESULTS: Significant clinical improvement was observed for mean ACQ scores from pre- to post-treatment (mean decrease 0.73, 95% CI 0.26-1.09, Cohen d = 2.25, p =.0014). No significant improvement was observed in the ACT, AQLQ, or spirometry, although inhaler use decreased for half of the subjects and did not change for the remaining subjects (i.e. none increased). Chest wall excursion and all ROM tests trended toward improvement, but was only statistically significant (p <.05) for the arm raise in the frontal plane. CONCLUSION: RFST may be a beneficial, nonpharmacological method to decrease asthma symptoms. Future studies should be conducted involving a larger sample size, longer intervention time, control group, and blood collection to test inflammatory mediators.

Stretching Impacts Inflammation Resolution in Connective Tissue.

Acute inflammation is accompanied from its outset by the release of specialized pro-resolving mediators (SPMs), including resolvins, that orchestrate the resolution of local inflammation. We showed earlier that, in rats with subcutaneous inflammation of the back induced by carrageenan, stretching for 10 min twice daily reduced inflammation and improved pain, 2 weeks after carrageenan injection. In this study, we hypothesized that stretching of connective tissue activates local pro-resolving mechanisms within the tissue in the acute phase of inflammation. In rats injected with carrageenan and randomized to stretch versus no stretch for 48 h, stretching reduced inflammatory lesion thickness and neutrophil count, and increased resolvin (RvD1) concentrations within lesions. Furthermore, subcutaneous resolvin injection mimicked the effect of stretching. In ex vivo experiments, stretching of connective tissue reduced the migration of neutrophils and increased tissue RvD1 concentration. These results demonstrate a direct mechanical impact of stretching on inflammation-regulation mechanisms within connective tissue.

Acupuncture, connective tissue, and peripheral sensory modulation.

Although considerable controversy surrounds the legitimacy of acupuncture as a treatment, a growing literature on the physiological effects of acupuncture needling in animals and humans is providing new insights into basic cellular mechanisms including connective tissue mechanotransduction and purinergic signaling. This review summarizes these findings and proposes a model combining connective tissue plasticity and peripheral sensory modulation in response to the sustained stretching of tissue that results from acupuncture needle manipulation.

Integrated multicomponent interventions to support healthy aging of the whole person.

Healthy aging is an integrated "whole person" process that involves an individual's biology, behavior, and social/physical environment. With the recent development of antiaging drugs, careful consideration of the respective roles of pharmacologic and nonpharmacologic approaches to both health and aging is in order. Recent advances in understanding the cellular and molecular mechanisms of aging are providing new measures that can be used as clinical outcomes in studying the impact of antiaging interventions in humans. This paper outlines the strategic interest of the National Center for Complementary and Integrative Health (NCCIH) in supporting the development, testing, and implementation of effective, scalable, and integrated multicomponent interventions to support healthy aging of the whole person.

Unmet Need: Mechanistic and Translational Studies of Sickle Cell Disease Pain as a Whole-Person Health Challenge.

Sickle cell disease (SCD) is a lifelong monogenic, autosomal-recessive blood disease that predominantly affects individuals of African descent and those who self-identify as Black or Hispanic. Common SCD pathophysiological processes include adhesion, hemolysis, hypoxia, ischemia, oxidative stress, and vaso-occlusion, which often lead to substantial comorbidities and complications. Pain is one of the most common and significant clinical complications for individuals with SCD. Despite advancements in understanding the pathophysiology of SCD, the ways in which SCD pathophysiological processes contribute to nociception and pain signaling, processing, and perception remain largely unclear. Pain management for individuals with SCD is complex and presents unique challenges that must be considered depending on the presenting pain type (eg, acute pain episode vs chronic pain). Racism, stigma (including stigma associated with opioid use), and limited resources present additional challenges. Limited research has been conducted on major clinical features of SCD pain such as its ischemic, inflammatory, and neuropathic components; on its transition from acute to chronic form and across the lifespan; and on factors influencing SCD pain perception. Research on and management of SCD pain requires a whole-person approach, bringing together investigators from multiple disciplines such as hematologists, organ biologists, pain experts, physiologists, neuroscientists, psychologists, geneticists, microbiologists, immunologists, behavioral scientists, and clinicians. Multidisciplinary cross-training, with different platforms for information dissemination and communication, could help promote basic, mechanistic, and translational research to inform the optimization of current treatment strategies and the development of novel therapies for SCD pain. PERSPECTIVE: This review presents the research challenges and negative impact of SCD pain, a grossly understudied condition in a highly underserved population. It also highlights the barriers and opportunities in SCD pain research and could help clinicians better understand current treatment strategies from the whole-person perspective.

Stress and matrix-responsive cytoskeletal remodeling in fibroblasts.

In areolar "loose" connective tissue, fibroblasts remodel their cytoskeleton within minutes in response to static stretch resulting in increased cell body cross-sectional area that relaxes the tissue to a lower state of resting tension. It remains unknown whether the loosely arranged collagen matrix, characteristic of areolar connective tissue, is required for this cytoskeletal response to occur. The purpose of this study was to evaluate cytoskeletal remodeling of fibroblasts in, and dissociated from, areolar and dense connective tissue in response to 2 h of static stretch in both native tissue and collagen gels of varying crosslinking. Rheometric testing indicated that the areolar connective tissue had a lower dynamic modulus and was more viscous than the dense connective tissue. In response to stretch, cells within the more compliant areolar connective tissue adopted a large "sheet-like" morphology that was in contrast to the smaller dendritic morphology in the dense connective tissue. By adjusting the in vitro collagen crosslinking, and the resulting dynamic modulus, it was demonstrated that cells dissociated from dense connective tissue are capable of responding when seeded into a compliant matrix, while cells dissociated from areolar connective tissue can lose their ability to respond when their matrix becomes stiffer. This set of experiments indicated stretch-induced fibroblast expansion was dependent on the distinct matrix material properties of areolar connective tissues as opposed to the cells' tissue of origin. These results also suggest that disease and pathological processes with increased crosslinks, such as diabetes and fibrosis, could impair fibroblast responsiveness in connective tissues.

Fibroblast cytoskeletal remodeling induced by tissue stretch involves ATP signaling.

Fibroblasts in whole areolar connective tissue respond to static stretching of the tissue by expanding and remodeling their cytoskeleton within minutes both ex vivo and in vivo. This study tested the hypothesis that the mechanism of fibroblast expansion in response to tissue stretch involves extracellular ATP signaling. In response to tissue stretch ex vivo, ATP levels in the bath solution increased significantly, and this increase was sustained for 20 min, returning to baseline at 60 min. No increase in ATP was observed in tissue incubated without stretch or tissue stretched in the presence of the Rho kinase inhibitor Y27632. The increase in fibroblast cross sectional area in response to tissue stretch was blocked by both suramin (a purinergic receptor blocker) and apyrase (an enzyme that selectively degrades extracellular ATP). Furthermore, connexin channel blockers (octanol and carbenoxolone), but not VRAC (fluoxetine) or pannexin (probenecid) channel blockers, inhibited fibroblast expansion. Together, these results support a mechanism in which extracellular ATP signaling via connexin hemichannels mediate the active change in fibroblast shape that occurs in response to a static increase in tissue length.

Cellular control of connective tissue matrix tension.

The biomechanical behavior of connective tissue in response to stretching is generally attributed to the molecular composition and organization of its extracellular matrix. It also is becoming apparent that fibroblasts play an active role in regulating connective tissue tension. In response to static stretching of the tissue, fibroblasts expand within minutes by actively remodeling their cytoskeleton. This dynamic change in fibroblast shape contributes to the drop in tissue tension that occurs during viscoelastic relaxation. We propose that this response of fibroblasts plays a role in regulating extracellular fluid flow into the tissue, and protects against swelling when the matrix is stretched. This article reviews the evidence supporting possible mechanisms underlying this response including autocrine purinergic signaling. We also discuss fibroblast regulation of connective tissue tension with respect to lymphatic flow, immune function, and cancer.

Spatial organization of fibroblast nuclear chromocenters: component tree analysis.

The nuclei of mouse connective tissue fibroblasts contain chromocenters which are well-defined zones of heterochromatin that can be used as positional landmarks to examine nuclear remodeling in response to a mechanical perturbation. This study used component tree analysis, an image segmentation algorithm that detects high intensity voxels that are topologically connected, to quantify the spatial organization of chromocenters in fibroblasts within whole mouse connective tissue fixed and stained with 4',6-diamidino-2-phenylindole (DAPI). The component tree analysis method was applied to confocal microscopy images of whole mouse areolar connective tissue incubated for 30 min ex vivo with or without static stretch. In stretched tissue, the mean distance between chromocenters within fibroblast nuclei was significantly greater (vs. non-stretched, P < 0.001), corresponding to an average of a 500-nm increase in chromocenter separation (~10% strain). There was no significant difference in chromocenter number or average size between stretch and no stretch. Average chromocenter distance was positively correlated with nuclear cross-sectional area (r = 0.78, P < 0.0001), and nuclear volume (r = 0.42, P < 0.0001), and negatively correlated with nuclear aspect ratio (r = -0.65, P < 0.0001) and nuclear concavity index (r = -0.44, P < 0.0001). These results demonstrate that component trees can be successfully applied to the morphometric analysis of nuclear chromocenters in fibroblasts within whole connective tissue. Static stretching of mouse areolar connective tissue for 30 min resulted in substantially increased separation of nuclear chromocenters in connective tissue fibroblasts. This interior remodeling of the nucleus induced by tissue stretch may impact transcriptionally active euchromatin within the inter-chromocenter space.

Total Force Fitness: Making Holistic, Integrated Whole-Person Research a DoD Priority.

Total Force Fitness (TFF) was conceived as a holistic framework for building and sustaining Human Performance Optimization for Warfighters and their families. As such, TFF research must also be holistic in nature. During the research breakout, group barriers and challenges to TFF research were discussed, and critical research focus areas were prioritized. The top approaches discussed were (1) using big data to identify best practices and health trajectories; (2) applying community-based participatory research principles to military units; (3) focusing on "Whole-Person," integrative research (physical, behavioral, spiritual, and biological) across the Department of Defense; and, finally, (4) prioritizing key opportunities to advance TFF across the active duty and Reserve/Guard enterprises and their families. The research group noted that coordinated action would be needed to move the prioritized agenda forward. Finally, translating research into action is essential because TFF is a way of honoring our service members as whole persons with careers, goals, and families.

Complementary and integrative health therapies in whole person resilience research.

Complementary and integrative health approaches can improve health and well-being, as well as play an important role in disease prevention. The concept of whole person health builds on these concepts by empowering individuals, families, communities, and populations to improve their health in multiple interconnected domains: biological, behavioural, social, and environmental. Research on whole person health involves studies of interconnected biological systems and complex approaches to prevention and treatment. Some of these approaches may involve methods of diagnosis and therapy that differ from those used in conventional Western medicine. Of growing interest is how complementary, integrative, and whole person health approaches contribute to resilience. This brief commentary describes an integrated framework for mapping the connections between various complementary and integrative health therapeutic inputs onto aspects of resilience, including the ability to resist, recover (partially or fully), adapt, and/or grow in response to a following a stressor. The authors present selected examples of research studies supported by the National Institutes of Health that test whether complementary and integrative health approaches can promote some aspect of resilience. We conclude with a discussion of the challenges and opportunities in incorporating the study of resilience in complementary, integrative, and whole person health research.

Live free or die: stretch-induced apoptosis occurs when adaptive reorientation of annulus fibrosus cells is restricted.

High matrix strains in the intervertebral disc occur during physiological motions and are amplified around structural defects in the annulus fibrosus (AF). It remains unknown if large matrix strains in the human AF result in localized cell death. This study investigated strain amplitudes and substrate conditions where AF cells were vulnerable to stretch-induced apoptosis. Human degenerated AF cells were subjected to 1 Hz-cyclic tensile strains for 24h on uniformly collagen coated substrates and on substrates with 40 µm stripes of collagen that restricted cellular reorientation. AF cells were capable of responding to stretch (stress fibers and focal adhesions aligned perpendicular to the direction of stretch), but were vulnerable to stretch-induced apoptosis when cytoskeletal reorientation was restricted, as could occur in degenerated states due to fibrosis and crosslink accumulation and at areas where high strains occur (around structural defects, delaminations, and herniations).

Making Connections to Improve Health Outcomes.

This issue of Global Advances in Health and Medicine sends a vital message about the importance of whole person health. Whole person health rests on the idea that our health involves multiple interconnected factors across physiological systems, as well as biological, behavioral, social, and environmental domains. The urgency of better understanding whole person health is highlighted by the current global health crisis. Yet, biomedical research often favors a reductionist approach. The current emphasis on diseases or single organ systems can fall short when it comes to addressing the interconnected factors that contribute to worse health outcomes. This, coupled with a fragmented health care delivery system, contributes to the challenges that patients face every day in becoming healthier. As part of the U.S. National Institutes of Health, our role at the National Center for Complementary and Integrative Health (NCCIH) is to foster research in this field. NCCIH's twenty years of research has built a body of knowledge that has established a clear path forward for exploring whole person health in the coming years. Within the framework of our strategic plan, NCCIH is working to build research methods for studying whole person health and explore how this understanding of health can transform the way complementary and integrative health is perceived and implemented within the wider health care delivery system.The collection of papers highlighted in this month's issue of Global Advances in Health and Medicine sends an important and encouraging signal about the efforts being made to deliver health care in a way that recognizes the importance of whole person health. Each of these studies provides new insights on how stakeholders might approach transforming the delivery of health care, integrating approaches that can improve health outcomes for people.

Fibroblast cytoskeletal remodeling contributes to connective tissue tension.

The visco-elastic behavior of connective tissue is generally attributed to the material properties of the extracellular matrix rather than cellular activity. We have previously shown that fibroblasts within areolar connective tissue exhibit dynamic cytoskeletal remodeling within minutes in response to tissue stretch ex vivo and in vivo. Here, we tested the hypothesis that fibroblasts, through this cytoskeletal remodeling, actively contribute to the visco-elastic behavior of the whole tissue. We measured significantly increased tissue tension when cellular function was broadly inhibited by sodium azide and when cytoskeletal dynamics were compromised by disrupting microtubules (with colchicine) or actomyosin contractility (via Rho kinase inhibition). These treatments led to a decrease in cell body cross-sectional area and cell field perimeter (obtained by joining the end of all of a fibroblast's processes). Suppressing lamellipodia formation by inhibiting Rac-1 decreased cell body cross-sectional area but did not affect cell field perimeter or tissue tension. Thus, by changing shape, fibroblasts can dynamically modulate the visco-elastic behavior of areolar connective tissue through Rho-dependent cytoskeletal mechanisms. These results have broad implications for our understanding of the dynamic interplay of forces between fibroblasts and their surrounding matrix, as well as for the neural, vascular, and immune cell populations residing within connective tissue.

Sensory innervation of the nonspecialized connective tissues in the low back of the rat.

Chronic musculoskeletal pain, including low back pain, is a worldwide debilitating condition; however, the mechanisms that underlie its development remain poorly understood. Pathological neuroplastic changes in the sensory innervation of connective tissue may contribute to the development of nonspecific chronic low back pain. Progress in understanding such potentially important abnormalities is hampered by limited knowledge of connective tissue's normal sensory innervation. The goal of this study was to evaluate and quantify the sensory nerve fibers terminating within the nonspecialized connective tissues in the low back of the rat. With 3-dimensional reconstructions of thick (30-80 µm) tissue sections we have for the first time conclusively identified sensory nerve fiber terminations within the collagen matrix of connective tissue in the low back. Using dye labeling techniques with Fast Blue, presumptive dorsal root ganglia cells that innervate the low back were identified. Of the Fast Blue-labeled cells, 60-88% also expressed calcitonin gene-related peptide (CGRP) immunoreactivity. Based on the immunolabeling with CGRP and the approximate size of these nerve fibers (≤2 µm) we hypothesize that they are Aδ or C fibers and thus may play a role in the development of chronic pain.

Paradoxes in acupuncture research: strategies for moving forward.

In November 2007, the Society for Acupuncture Research (SAR) held an international symposium to mark the 10th anniversary of the 1997 NIH Consensus Development Conference on Acupuncture. The symposium presentations revealed the considerable maturation of the field of acupuncture research, yet two provocative paradoxes emerged. First, a number of well-designed clinical trials have reported that true acupuncture is superior to usual care, but does not significantly outperform sham acupuncture, findings apparently at odds with traditional theories regarding acupuncture point specificity. Second, although many studies using animal and human experimental models have reported physiological effects that vary as a function of needling parameters (e.g., mode of stimulation) the extent to which these parameters influence therapeutic outcomes in clinical trials is unclear. This White Paper, collaboratively written by the SAR Board of Directors, identifies gaps in knowledge underlying the paradoxes and proposes strategies for their resolution through translational research. We recommend that acupuncture treatments should be studied (1) "top down" as multi-component "whole-system" interventions and (2) "bottom up" as mechanistic studies that focus on understanding how individual treatment components interact and translate into clinical and physiological outcomes. Such a strategy, incorporating considerations of efficacy, effectiveness and qualitative measures, will strengthen the evidence base for such complex interventions as acupuncture.

Reduced thoracolumbar fascia shear strain in human chronic low back pain.

BACKGROUND: The role played by the thoracolumbar fascia in chronic low back pain (LBP) is poorly understood. The thoracolumbar fascia is composed of dense connective tissue layers separated by layers of loose connective tissue that normally allow the dense layers to glide past one another during trunk motion. The goal of this study was to quantify shear plane motion within the thoracolumbar fascia using ultrasound elasticity imaging in human subjects with and without chronic low back pain (LBP). METHODS: We tested 121 human subjects, 50 without LBP and 71 with LBP of greater than 12 months duration. In each subject, an ultrasound cine-recording was acquired on the right and left sides of the back during passive trunk flexion using a motorized articulated table with the hinge point of the table at L4-5 and the ultrasound probe located longitudinally 2 cm lateral to the midline at the level of the L2-3 interspace. Tissue displacement within the thoracolumbar fascia was calculated using cross correlation techniques and shear strain was derived from this displacement data. Additional measures included standard range of motion and physical performance evaluations as well as ultrasound measurement of perimuscular connective tissue thickness and echogenicity. RESULTS: Thoracolumbar fascia shear strain was reduced in the LBP group compared with the No-LBP group (56.4% ± 3.1% vs. 70.2% ± 3.6% respectively, p < .01). There was no evidence that this difference was sex-specific (group by sex interaction p = .09), although overall, males had significantly lower shear strain than females (p = .02). Significant correlations were found in male subjects between thoracolumbar fascia shear strain and the following variables: perimuscular connective tissue thickness (r = -0.45, p <.001), echogenicity (r = -0.28, p < .05), trunk flexion range of motion (r = 0.36, p < .01), trunk extension range of motion (r = 0.41, p < .01), repeated forward bend task duration (r = -0.54, p < .0001) and repeated sit-to-stand task duration (r = -0.45, p < .001). CONCLUSION: Thoracolumbar fascia shear strain was ~20% lower in human subjects with chronic low back pain. This reduction of shear plane motion may be due to abnormal trunk movement patterns and/or intrinsic connective tissue pathology. There appears to be some sex-related differences in thoracolumbar fascia shear strain that may also play a role in altered connective tissue function.

Reconnecting the Brain With the Rest of the Body in Musculoskeletal Pain Research.

A challenge in understanding chronic musculoskeletal pain is that research is often siloed between neuroscience, physical therapy/rehabilitation, orthopedics, and rheumatology which focus respectively on 1) neurally mediated effects on pain processes, 2) behavior and muscle activity, 3) tissue structure, and 4) inflammatory processes. Although these disciplines individually study important aspects of pain, there is a need for more cross-disciplinary research that can bridge between them. Identifying the gaps in knowledge is important to understand the whole body, especially at the interfaces between the silos-between brain function and behavior, between behavior and tissue structure, between musculoskeletal and immune systems, and between peripheral tissues and the nervous system. Research on "mind and body" practices can bridge across these silos and encourage a "whole person" approach to better understand musculoskeletal pain by bringing together the brain and the rest of the body. PERSPECTIVE: Research on chronic musculoskeletal pain is limited by significant knowledge gaps. To be fully integrated, musculoskeletal pain research will need to bridge across tissues, anatomical areas, and body systems. Research on mind and body approaches encourages a "whole person" approach to better understand musculoskeletal pain.

Fascia Mobility, Proprioception, and Myofascial Pain.

The network of fasciae is an important part of the musculoskeletal system that is often overlooked. Fascia mobility, especially along shear planes separating muscles, is critical for musculoskeletal function and may play an important, but little studied, role in proprioception. Fasciae, especially the deep epimysium and aponeuroses, have recently been recognized as highly innervated with small diameter fibers that can transmit nociceptive signals, especially in the presence of inflammation. Patients with connective tissue hyper- and hypo-mobility disorders suffer in large number from musculoskeletal pain, and many have abnormal proprioception. The relationships among fascia mobility, proprioception, and myofascial pain are largely unstudied, but a better understanding of these areas could result in improved care for many patients with musculoskeletal pain.