Although stem cell therapy has tremendous therapeutic potential, clinical translation of stem cell therapy has yet to be fully realized. Recently, patient comorbidities and lifestyle choices have emerged to be important factors in the efficacy of stem cell therapy. Tobacco usage is an important risk factor for numerous diseases, and nicotine exposure specifically has become increasing more prevalent with the rising use of electronic cigarettes. This review describes the effects of nicotine exposure on the function of various stem cells. We place emphasis on the differential effects of nicotine exposure in vitro and as well as in preclinical models. Further research on the effects of nicotine on stem cells will deepen our understanding of how lifestyle choices can impact the outcome of stem cell therapies.

Down syndrome cell adhesion molecules (DSCAMs) are broadly expressed in nervous systems and play conserved roles in programmed cell death, neuronal migration, axon guidance, neurite branching and spacing, and synaptic targeting. However, DSCAMs appear to have distinct functions in different vertebrate animals, and little is known about their functions outside the retina. We leveraged the genetic tractability and optical accessibility of larval zebrafish to investigate the expression and function of a DSCAM family member, dscamb. Using targeted genome editing to create transgenic reporters and loss-of-function mutant alleles, we discovered that dscamb is expressed broadly throughout the brain, spinal cord, and peripheral nervous system, but is not required for overall structural organization of the brain. Despite the absence of obvious anatomical defects, homozygous dscamb mutants were deficient in their ability to ingest food and rarely survived to adulthood. Thus, we have discovered a novel function for dscamb in feeding behavior. The mutant and transgenic lines generated in these studies will provide valuable tools for identifying the molecular and cellular bases of these behaviors.

Wound healing is a complex process that involves sequential phases that overlap in time and space and affect each other dynamically at the gene and protein levels. We previously showed that insulin accelerates wound healing by stimulating faster and regenerative healing. One of the processes that insulin stimulates is an increase in monocyte/macrophage chemotaxis. In this study, we performed experiments in vivo and in vitro to elucidate the signaling transduction pathways that are involved in insulin-induced monocyte/macrophage chemotaxis. We found that insulin stimulates THP-1 cell chemotaxis in a dose-dependent and insulin receptor-dependent manner. We also show that the kinases PI3K-Akt, SPAK/JNK, and p38 MAPK are key molecules in the insulin-induced signaling pathways that lead to chemoattraction of the THP-1 cell. Furthermore, both PI3K-Akt and SPAK/JNK signaling involve Rac1 activation, an important molecule in regulating cell motility. Indeed, topical application of Rac1 inhibitor at an early stage during the healing process caused delayed and impaired healing even in the presence of insulin. These results delineate cell and molecular mechanisms involved in insulin-induced chemotaxis of monocyte/macrophage, cells that are critical for proper healing.

Antimicrobial peptides (AMPs) are good candidates to treat burn wounds, a major cause of morbidity, impaired life quality and resources consumption in developed countries. We took advantage of a commercially available hydrogel, Carbopol®, a vehicle for topical administration that maintains a moist environment within the wound site. We hypothesized that the incorporation of LLKKK18 conjugated to dextrin would improve the healing process in rat burns. Whereas the hydrogel improves healing, LLKKK18 released from the dextrin conjugates further accelerated wound closure, and simultaneously improving the quality of healing. Indeed, the release of LLKKK18 reduced oxidative stress and inflammation (low neutrophil and macrophage infiltration and pro-inflammatory cytokines levels). Importantly, it induced a faster resolution of the inflammatory stage through early M2 macrophage recruitment. In addition, LLKKK18 stimulated angiogenesis (increased VEGF and microvessel development in vivo). Moreover, collagen staining evaluated by Masson's Trichrome was visually much more intense after treatment with LLKKK18, suggesting higher collagen deposition. Overall, we generated an effective, safe and inexpensive formulation that maintains a moist environment in the wound, easy to apply and remove, and with potential to prevent infection due to the presence of an antimicrobial peptide. These findings propel us to further study this LLKKK18-containing formulation, setting the foundations towards a potential therapeutic approach for burn wound treatment.

Statement of significance

This work presents a newly developed formulation that holds great potential as a therapeutic approach for burn treatment. It is based on the sustained delivery of an antimicrobial peptide - LLKKK18 - from conjugates with dextrin, after degradation of dextrin backbone upon exposure to wound α-amylases. Conjugates were further embedded in Carbopol®, a commercially available hydrogel, suitable for topical administration and that provides a moist environment to the wound. Overall, we obtained an efficient, safe and non-expensive formulation that improves burn wound healing, maintains a moist environment within the wound, is easy to apply-and-remove, and has potential to prevent infection due to the presence of an antimicrobial peptide. Importantly, this is the first time the wound healing ability of LLKKK18 is demonstrated and that its main mechanisms of action are identified.

Volumetric muscle loss (VML) is associated with irreversibly impaired muscle function due to traumatic injury. Experimental approaches to treat VML include the delivery of basic fibroblast growth factor (bFGF) or rehabilitative exercise. The objective of this study was to compare the effects of spatially nanopatterned collagen scaffold implants with either bFGF delivery or in conjunction with voluntary exercise. Aligned nanofibrillar collagen scaffold bundles were adsorbed with bFGF, and the bioactivity of bFGF-laden scaffolds was examined by skeletal myoblast or endothelial cell proliferation. The therapeutic efficacy of scaffold implants with either bFGF release or exercise was examined in a murine VML model. Our results show an initial burst release of bFGF from the scaffolds, followed by a slower release over 21 days. The released bFGF induced myoblast and endothelial cell proliferation in vitro. After 3 weeks of implantation in a mouse VML model, twitch force generation was significantly higher in mice treated with bFGF-laden scaffolds compared to bFGF-laden scaffolds with exercise. However, myofiber density was not significantly improved with bFGF scaffolds or voluntary exercise. In contrast, the scaffold implant with exercise induced more re-innervation than all other groups. These results highlight the differential effects of bFGF and exercise on muscle regeneration.

Objective

Lifestyle choices such as tobacco and e-cigarette use are a risk factor for peripheral arterial disease (PAD) and may influence therapeutic outcomes. The effect of chronic nicotine exposure on the angiogenic capacity of human induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) was assessed in a murine model of PAD.

Methods

Mice were exposed to nicotine or phosphate-buffered saline (PBS) for 28 days, followed by induction of limb ischemia and iPSC-EC transplantation. Cells were injected into the ischemic limb immediately after induction of hindlimb ischemia and again 7 days later. Limb perfusion was assessed by laser Doppler spectroscopy, and transplant cell survival was monitored for 14 days afterward using bioluminescence imaging, followed by histological analysis of angiogenesis.

Results

Transplant cell retention progressively decreased over time after implantation based on bioluminescence imaging, and there were no significant differences in cell survival between mice with chronic exposure to nicotine or PBS. However, compared with mice without nicotine exposure, mice with prior nicotine exposure had had an impaired therapeutic response to iPSC-EC therapy based on decreased vascular perfusion recovery. Mice with nicotine exposure, followed by cell transplantation, had significantly lower mean perfusion ratio after 14 days (0.47 ± 0.07) compared with mice undergoing cell transplantation without prior nicotine exposure (0.79 ± 0.11). This finding was further supported by histological analysis of capillary density, in which animals with prior nicotine exposure had a lower capillary density (45.9 ± 4.7 per mm²) compared with mice without nicotine exposure (66.5 ± 8.1 per mm²). Importantly, the ischemic limbs mice exposed to nicotine without cell therapy also showed significant impairment in perfusion recovery after 14 days, compared with mice that received PBS + iPSC-EC treatment. This result suggested that mice without chronic nicotine exposure could respond to iPSC-EC implantation into the ischemic limb by inducing perfusion recovery, whereas mice with chronic nicotine exposure did not respond to iPSC-EC therapy.

Conclusions

Together, these findings show that chronic nicotine exposure adversely affects the ability of iPSC-EC therapy to promote vascular perfusion recovery and angiogenesis in a murine PAD model.

Third hand smoke (THS) is the accumulation of second hand smoke (SHS) toxins on surfaces in homes, cars, clothing and hair of smokers. It is known that 88M US nonsmokers ≥3 years old living in homes of smokers are exposed to THS toxicants and show blood cotinine levels of ≥0.05 ng/ml, indicating that the toxins are circulating in their circulatory systems. The goal of the present study is to investigate the mechanisms by which THS causes impaired wound healing. We show that mice living under conditions that mimic THS exposure in humans display delayed wound closure, impaired collagen deposition, altered inflammatory response, decreased angiogenesis, microvessels with fibrin cuffs and a highly proteolytic wound environment. Moreover, THS-exposed mouse wounds have high levels of oxidative stress and significantly lower levels of antioxidant activity leading to molecular damage, including protein nitration, lipid peroxidation and DNA damage that contribute to tissue dysfunction. Furthermore, we show that elastase is elevated, suggesting that elastin is degraded and the plasticity of the wound tissue is decreased. Taken together, our results lead us to conclude that THS toxicants delay and impair wound healing by disrupting the sequential processes that lead to normal healing. In addition, the lack of elastin results in loss of wound plasticity, which may be responsible for reopening of wounds.

Burn wound healing involves a complex set of overlapping processes in an environment conducive to ischaemia, inflammation and infection costing $7.5 billion/year in the U.S.A. alone, in addition to the morbidity and mortality that occur when the burns are extensive. We previously showed that insulin, when topically applied to skin excision wounds, accelerates re-epithelialization and stimulates angiogenesis. More recently, we developed an alginate sponge dressing (ASD) containing insulin encapsulated in PLGA [poly(D,L-lactic-co-glycolic acid)] microparticles that provides a sustained release of bioactive insulin for >20 days in a moist and protective environment. We hypothesized that insulin-containing ASD accelerates burn healing and stimulates a more regenerative, less scarring healing. Using heat-induced burn injury in rats, we show that burns treated with dressings containing 0.04 mg insulin/cm(2) every 3 days for 9 days have faster closure, a higher rate of disintegration of dead tissue and decreased oxidative stress. In addition, in insulin-treated wounds, the pattern of neutrophil inflammatory response suggests faster clearing of the burned dead tissue. We also observe faster resolution of the pro-inflammatory macrophages. We also found that insulin stimulates collagen deposition and maturation with the fibres organized more like a basket weave (normal skin) than aligned and cross-linked (scar tissue). In summary, application of ASD-containing insulin-loaded PLGA particles on burns every 3 days stimulates faster and more regenerative healing. These results suggest insulin as a potential therapeutic agent in burn healing and, because of its long history of safe use in humans, insulin could become one of the treatments of choice when repair and regeneration are critical for proper tissue function.

Abdominal aortic aneurysm (AAA) is associated with the loss of vascular smooth muscle cells (SMCs) within the vessel wall. Direct delivery of therapeutic cells is challenging due to impaired mechanical integrity of the vessel wall. We hypothesized that porous collagen scaffolds can be an effective vehicle for the delivery of human-derived SMCs to the site of AAA. The purpose was to evaluate if the delivery of cell-seeded scaffolds can abrogate progressive expansion in a mouse model of AAA. Collagen scaffolds seeded with either primary human aortic SMCs or induced pluripotent stem cell derived-smooth muscle progenitor cells (iPSC-SMPs) had >80% in vitro cell viability and >75% cell penetrance through the scaffold's depth, while preserving smooth muscle phenotype. The cell-seeded scaffolds were successfully transplanted onto the murine aneurysm peri-adventitia on day 7 following AAA induction using pancreatic porcine elastase infusion. Ultrasound imaging revealed that SMC-seeded scaffolds significantly reduced the aortic diameter by 28 days, compared to scaffolds seeded with iPSC-SMPs or without cells (acellular scaffold), respectively. Bioluminescence imaging demonstrated that both cell-seeded scaffold groups had cellular localization to the aneurysm but a decline in survival with time. Histological analysis revealed that both cell-seeded scaffold groups had more SMC retention and less macrophage invasion into the medial layer of AAA lesions, when compared to the acellular scaffold treatment group. Our data suggest that scaffold-based SMC delivery is feasible and may constitute a platform for cell-based AAA therapy.

Undergraduate students participating in the UCLA Undergraduate Research Consortium for Functional Genomics (URCFG) have conducted a two-phased screen using RNA interference (RNAi) in combination with fluorescent reporter proteins to identify genes important for hematopoiesis in Drosophila. This screen disrupted the function of approximately 3500 genes and identified 137 candidate genes for which loss of function leads to observable changes in the hematopoietic development. Targeting RNAi to maturing, progenitor, and regulatory cell types identified key subsets that either limit or promote blood cell maturation. Bioinformatic analysis reveals gene enrichment in several previously uncharacterized areas, including RNA processing and export and vesicular trafficking. Lastly, the participation of students in this course-based undergraduate research experience (CURE) correlated with increased learning gains across several areas, as well as increased STEM retention, indicating that authentic, student-driven research in the form of a CURE represents an impactful and enriching pedagogical approach.