Development of DNA Based Active Macro–Materials for Biology and Medicine: A Review

DNA was first discovered as the carrier of genetic information for the majority of the known living organisms, encoding the secret of life. Its delicate design based upon double helical structure and base pairing offers a stable and reliable media for storing hereditary codes, laying the foundation for the central dogma (Watson et al. 2003). The impact of this molecule is far reaching into scientific community and our society, as manifested in many fields, for instance, forensics (Budowle et al. 2003), besides medicine. To date, a great deal of research effort has been directed towards understandin

The Role of Macrophages in Acute and Chronic Wound Healing and Interventions to Promote Pro-wound Healing Phenotypes

Macrophages play key roles in all phases of adult wound healing, which are inflammation, proliferation, and remodeling. As wounds heal, the local macrophage population transitions from predominantly pro-inflammatory (M1-like phenotypes) to anti-inflammatory (M2-like phenotypes). Non-healing chronic wounds, such as pressure, arterial, venous, and diabetic ulcers indefinitely remain in inflammation—the first stage of wound healing. Thus, local macrophages retain pro-inflammatory characteristics. This review discusses the physiology of monocytes and macrophages in acute wound healing and the different phenotypes described in the literature for both in vitro and in vivo models. We also discuss aberrations that occur in macrophage populations in chronic wounds, and attempts to restore macrophage function by therapeutic approaches. These include endogenous M1 attenuation, exogenous M2 supplementation and endogenous macrophage modulation/M2 promotion via mesenchymal stem cells, growth factors, biomaterials, heme oxygenase-1 (HO-1) expression, and oxygen therapy. We recognize the challenges and controversies that exist in this field, such as standardization of macrophage phenotype nomenclature, definition of their distinct roles and understanding which phenotype is optimal in order to promote healing in chronic wounds

Prostaglandin E2 produced by alginate-encapsulated mesenchymal stromal cells modulates the astrocyte inflammatory response

Astroglia are well known for their role in propagating secondary injury following brain trauma. Modulation of this injury cascade, including inflammation, is essential to repair and recovery. Mesenchymal stromal cells (MSCs) have been demonstrated as trophic mediators in several models of secondary CNS injury, however, there has been varied success with the use of direct implantation due to a failure to persist at the injury site. To achieve sustained therapeutic benefit, we have encapsulated MSCs in alginate microspheres and evaluated the ability of these encapsulated MSCs to attenuate neuro-inflammation. In this study, astroglial cultures were administered lipopolysaccharide (LPS) to induce inflammation and immediately co-cultured with encapsulated or monolayer human MSCs. Cultures were assayed for the pro-inflammatory cytokine tumor necrosis factor alpha (TNF-[Formula: see text] produced by astroglia, MSC-produced prostaglandin E2, and expression of neurotrophin-associated genes. We found that encapsulated MSCs significantly reduced TNF-[Formula: see text] produced by LPS-stimulated astrocytes more effectively than monolayer MSCs, and this enhanced benefit commences earlier than that of monolayer MSCs. Furthermore, in support of previous findings, encapsulated MSCs constitutively produced high levels of PGE2, while monolayer MSCs required the presence of inflammatory stimuli to induce PGE2production. The early, constitutive presence of PGE2significantly reduced astrocyte-produced TNF-[Formula: see text], while delayed administration had no effect. Finally, MSC-produced PGE2was not only capable of modulating inflammation, but appeared to have an additional role in stimulating astrocyte neurotrophin production. Overall, these results support the enhanced benefit of encapsulated MSC treatment, both in modulating the inflammatory response and providing neuroprotection.</jats:p

Status of the Archaeal and Bacterial Census: an Update

A census is typically carried out for people across a range of geographical levels; however, microbial ecologists have implemented a molecular census of bacteria and archaea by sequencing their 16S rRNA genes. We assessed how well the census of full-length 16S rRNA gene sequences is proceeding in the context of recent advances in high-throughput sequencing technologies because full-length sequences are typically used as references for classification of the short sequences generated by newer technologies. Among the 1,411,234 and 53,546 full-length bacterial and archaeal sequences, 94.5% and 95.1% of the bacterial and archaeal sequences, respectively, belonged to operational taxonomic units (OTUs) that have been observed more than once. Although these metrics suggest that the census is approaching completion, 29.2% of the bacterial and 38.5% of the archaeal OTUs have been observed more than once. Thus, there is still considerable diversity to be explored. Unfortunately, the rate of new full-length sequences has been declining, and new sequences are primarily being deposited by a small number of studies. Furthermore, sequences from soil and aquatic environments, which are known to be rich in bacterial diversity, represent only 7.8 and 16.5% of the census, while sequences associated with host-associated environments represent 55.0% of the census. Continued use of traditional approaches and new technologies such as single-cell genomics and short-read assembly are likely to improve our ability to sample rare OTUs if it is possible to overcome this sampling bias. The success of ongoing efforts to use short-read sequencing to characterize archaeal and bacterial communities requires that researchers strive to expand the depth and breadth of this census

An in vitro model of the macrophage-endothelial interface to characterize CAR T-cell induced cytokine storm

Abstract Chimeric Antigen Receptor (CAR) T-cell therapy is a highly effective treatment for B-cell malignancies but limited in use due to clinically significant hyperinflammatory toxicities. Understanding the pathophysiologic mechanisms which mediate these toxicities can help identify novel management strategies. Here we report a novel in vitro model of the macrophage-endothelial interface to study the effects of CAR T-cell-induced cytokine storm. Using this model, we demonstrate that macrophage-mediated inflammation is regulated by endothelial cell activity. Furthermore, endothelial inflammation occurs independently of macrophages following exposure to CAR T-cell products and the induced endothelial inflammation potentiates macrophage-mediated inflammatory signaling, leading to a hyperinflammatory environment. While corticosteroids, the current gold standard of care, attenuate the resulting macrophage inflammatory signaling, the endothelial activity remains refractory to this treatment strategy. Utilizing a network model, coupled to in vitro secretion profiling, we identified STAT3 programming as critical in regulating this endothelial behavior. Lastly, we demonstrate how targeting STAT3 activity can abrogate endothelial inflammation and attenuate this otherwise hyperinflammatory environment. Our results demonstrate that endothelial cells play a central role in the pathophysiology of CAR T-cell toxicities and targeting the mechanisms driving the endothelial response can guide future clinical management

The Relationship Between Fibroblast Growth and the Dynamic Stiffnesses of a DNA Crosslinked Hydrogel

The microenvironment of cells is dynamic and undergoes remodeling with time. This is evident in development, aging, pathological processes, and at tissue-biomaterial interfaces. But in contrast, the majority of the biomimetic materials have static properties. Here, we show that a previously developed DNA crosslinked hydrogel circumvents the need of environmental factors and undergoes controlled stiffness change via DNA delivery, a feasible approach to initiate property changes in vivo, different from previous attempts. Two types of fibroblasts, L929 and GFP, were subject to the alterations in substrate rigidity presented in the hydrogels. Our results show that exogenous DNA does not cause appreciable cell shape change. Cells do respond to mechanical alterations as demonstrated in the cell projection area and polarity (e.g., Soft vs. Soft → Medium), and the responses vary depending on magnitude (e.g., Soft → Medium vs. Soft → Stiff) and range of stiffness changes (e.g., Soft → Medium vs. Medium → Stiff). The two types of fibroblasts share specific responses in common (e.g., Soft → Medium), while differ in others (e.g., Medium → Stiff). For each cell type, the projection area and polarity respond differently. This approach provides insight into pathology (e.g., cancer) and tissue functioning, and assists in designing biomaterials with controlled dynamic stiffness by choosing the range and magnitude of stiffness change

Effect of Dynamic Stiffness of the Substrates on Neurite Outgrowth by Using a DNA-Crosslinked Hydrogel

Central nervous system tissues, like other tissue types, undergo constant remodeling, which potentially leads to changes in their mechanical stiffness. Moreover, mechanical compliance of central nervous system tissues can also be modified under external load such as that experienced in traumatic brain or spinal cord injury, and during pathological processes. Thus, the neuronal responses to the dynamic stiffness of the microenvironment are of significance. In this study, we induced decrease in stiffness by using a DNA-crosslinked hydrogel, and subjected rat spinal cord neurons to such dynamic stiffness. The neurons respond to the dynamic cues as evidenced by the primary neurite structure, and the response from each neurite property (e.g., axonal length and primary dendrite number) is consistent with the behavior on static gels of same substrate rigidity, with one exception of mean primary dendrite length. The results on cell population distribution confirm the neuronal responses to the dynamic stiffness. Quantification on the focal adhesion kinase expression in the neuronal cell body on dynamic gels suggests that neurons also modify adhesion in coping with the dynamic stiffnesses. The results reported here extend the neuronal mechanosensing capability to dynamic stiffness of extracellular matrix, and give rise to a novel way of engineering neurite outgrowth in time dimension