A human omentum-specific mesothelial-like stromal population inhibits adipogenesis through IGFBP2 secretion.

Adipose tissue plasticity is orchestrated by molecularly and functionally diverse cells within the stromal vascular fraction (SVF). Although several mouse and human adipose SVF cellular subpopulations have by now been identified, we still lack an understanding of the cellular and functional variability of adipose stem and progenitor cell (ASPC) populations across human fat depots. To address this, we performed single-cell and bulk RNA sequencing (RNA-seq) analyses of >30 SVF/Lin- samples across four human adipose depots, revealing two ubiquitous human ASPC (hASPC) subpopulations with distinct proliferative and adipogenic properties but also depot- and BMI-dependent proportions. Furthermore, we identified an omental-specific, high IGFBP2-expressing stromal population that transitions between mesothelial and mesenchymal cell states and inhibits hASPC adipogenesis through IGFBP2 secretion. Our analyses highlight the molecular and cellular uniqueness of different adipose niches, while our discovery of an anti-adipogenic IGFBP2+ omental-specific population provides a new rationale for the biomedically relevant, limited adipogenic capacity of omental hASPCs.

The Effect of Hybrosome (Umbilical Cord Blood Exosome-Liposome Hybrid Vesicles) on Human Dermal Cells In Vitro.

Background: Wound healing is a process that involves multiple physiological steps, and despite the availability of various wound treatment methods, their effectiveness is still limited due to several factors, including cost, efficiency, patient-specific requirements, and side effects. In recent years, nanovesicles called exosomes have gained increasing attention as a potential wound care solution due to their unique cargo components which enable cell-to-cell communication and regulate various biological processes. Umbilical cord blood plasma (UCBP) exosomes have shown promise in triggering beneficial signaling pathways that aid in cell proliferation and wound healing. However, there is still very limited information about the wound-healing effect of UCBP exosomes in the literature. Objectives: The primary objective of this study was to investigate the "hybrosome" technology generated with calf UCBP-derived exosome-liposome combination. Methods: The authors developed hybrosome technology by fusing cord blood exosome membranes with liposomes. Nanovesicle characterization, cell proliferation assay, wound-healing scratch assay, immunohistochemistry analysis, anti-inflammation assay, real-time polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay, and cellular uptake studies were performed using the novel hybrid exosomes. Results: Experimental results showed that hybrosome increases cell proliferation and migration by 40% to 50%, depending on the dose, and induces an anti-inflammatory effect on different cell lines as well as increased wound healing-related gene expression levels in dermal cells in vitro. All in all, this research widens the scope of wound-healing therapeutics to the novel hybrosome technology. Conclusions: UCBP-based applications have the potential for wound treatments and are promising in the development of novel therapies. This study shows that hybrosomes have outstanding abilities in wound healing using in vitro approaches.

Hybrid Stromal Vascular Fraction (Hybrid-SVF): A New Paradigm in Mechanical Regenerative Cell Processing.

Enzymatic digestion of extracellular matrix (ECM) from lipoaspirate is the conventional form of harvesting stromal vascular fraction (SVF) called enzymatically digested SVF (E-SVF). Mechanical SVF (M-SVF) isolation has emerged as an alternative method, but it has also some limitations in terms of lower cell viability and diminished cell counts. To enhance the SVF qualitatively and quantitatively, we propose a novel concept called "hybrid-SVF," in which we combine M-SVF with the concentrated parts of adipose tissue after centrifugation, which is called stromal vascular matrix (SVM). Methods: Hybrid-SVF injection was applied as an adjunctive therapy to fat grafting in 88 patients and 11 samples were evaluated in the laboratory for cell count, viability and cell activity. Results: Experimental results determined that SVM part showed higher cellular activity. SVM and M-SVF showed higher cellular potency than E-SVF. Clinically, none of the patients required an additional session for fat grafting since there was no significant graft resorption. However, seven patients asked for further volume augmentation due to their individual preferences. No major complication was encountered. Conclusions: The usage of hybrid-SVF has a very high regenerative potential due to the ECM support and exceptionally high cell yield in addition to preserved cell potency. Although there are ongoing studies focusing on optimizing cell counts and further clinical applications, we believe that our preliminary results might create a paradigm shift in the area of regenerative fat grafting.

Cellular Optimization of Nanofat: Comparison of Two Nanofat Processing Devices in Terms of Cell Count and Viability.

BACKGROUND: Nanofat was introduced by Tonnard and Verpaele in 2013. Their initial observations in intradermal applications showed improvement in the appearance of the skin. Since then, a number of Nanofat devices have been introduced. The cellular content in the processing of Nanofat is not the same in every device, yet the cellular composition is responsible for the biologic action of Nanofat. The authors sought to find a different means to produce a matrix rich Nanofat to optimize the cellular content. OBJECTIVES: The primary objective of this study was to compare cell counts, cultures, and cell viabilities produced by LipocubeNano (Lipocube, Inc., London, UK) in comparison to Tulip's NanoTransfer (Tulip Medical, San Diego, CA) processing methods. METHODS: Twenty milliliters of fat were harvested from 10 patients in order to test two methods of Nanofat production. Ten milliliters of fat were used to assess each method and, after the final product was obtained, enzymatic digestion for stromal vascular fraction (SVF) isolation was performed. A Muse Flow-cytometer was used to measure cell counts and cell viabilities, cell cultures were performed, and cell images were taken with a florescent microscope. RESULTS: The LipocubeNano was shown to be superior to Tulip's NanoTransfer system of progressive downsizing with final filtering, which appeared to trap more fibrous tissue leading to lower amounts of SVF. LipocubeNano resulted in higher cell counts (2.24 × 106/cc), whereas Tulip's NanoTransfer method resulted in a lower cell count at 1.44 × 106/cc. Cell viability was the same (96.05%) in both groups. CONCLUSIONS: Nanofat from LipocubeNano has a higher regenerative cell count and more SVF cells than the other common mechanical method of Nanofat processing. This new means of mechanical processing preserves more matrix, optimizing the cellular content of the Nanofat, thus having potentially a higher regenerative effect.

A stromal cell population that inhibits adipogenesis in mammalian fat depots.

Adipocyte development and differentiation have an important role in the aetiology of obesity and its co-morbidities1,2. Although multiple studies have investigated the adipogenic stem and precursor cells that give rise to mature adipocytes3-14, our understanding of their in vivo origin and properties is incomplete2,15,16. This is partially due to the highly heterogeneous and unstructured nature of adipose tissue depots17, which has proven difficult to molecularly dissect using classical approaches such as fluorescence-activated cell sorting and Cre-lox lines based on candidate marker genes16,18. Here, using the resolving power of single-cell transcriptomics19 in a mouse model, we reveal distinct subpopulations of adipose stem and precursor cells in the stromal vascular fraction of subcutaneous adipose tissue. We identify one of these subpopulations as CD142+ adipogenesis-regulatory cells, which can suppress adipocyte formation in vivo and in vitro in a paracrine manner. We show that adipogenesis-regulatory cells are refractory to adipogenesis and that they are functionally conserved in humans. Our findings point to a potentially critical role for adipogenesis-regulatory cells in modulating adipose tissue plasticity, which is linked to metabolic control, differential insulin sensitivity and type 2 diabetes.

Predictability of the individual clinical outcome of extracorporeal shock wave therapy for cellulite.

BACKGROUND: Extracorporeal shock wave therapy has been successfully introduced for the treatment of cellulite in recent years. However, it is still unknown whether the individual clinical outcome of cellulite treatment with extracorporeal shock wave therapy can be predicted by the patient's individual cellulite grade at baseline, individual patient age, body mass index (BMI), weight, and/or height. METHODS: Fourteen Caucasian females with cellulite were enrolled in a prospective, single-center, randomized, open-label Phase II study. The mean (± standard error of the mean) cellulite grade at baseline was 2.5±0.09 and mean BMI was 22.8±1.17. All patients were treated with radial extracorporeal shock waves using the Swiss DolorClast(®) device (Electro Medical Systems, S.A., Nyon, Switzerland). Patients were treated unilaterally with 2 weekly treatments for 4 weeks on a randomly selected side (left or right), totaling eight treatments on the selected side. Treatment was performed at 3.5-4.0 bar, with 15,000 impulses per session applied at 15 Hz. Impulses were homogeneously distributed over the posterior thigh and buttock area (resulting in 7,500 impulses per area). Treatment success was evaluated after the last treatment and 4 weeks later by clinical examination, photographic documentation, contact thermography, and patient satisfaction questionnaires. RESULTS: The mean cellulite grade improved from 2.5±0.09 at baseline to 1.57±0.18 after the last treatment (ie, mean δ-1 was 0.93 cellulite grades) and 1.68±0.16 at follow-up (ie, mean δ-2 was 0.82 cellulite grades). Compared with baseline, no patient's condition worsened, the treatment was well tolerated, and no unwanted side effects were observed. No statistically significant (ie, P<0.05) correlation was found between individual values for δ-1 and δ-2 and cellulite grade at baseline, BMI, weight, height, or age. CONCLUSION: Radial shock wave therapy is a safe and effective treatment option for cellulite. The individual clinical outcome cannot be predicted by the patient's individual cellulite grade at baseline, BMI, weight, height, or age.

Optimizing the arterialized venous flap.

BACKGROUND: Outcome of arterialized venous flaps is quite varied. The authors' initial experiments showed that a good vascular bed contributes significantly to survival of the flap. In continuation of these experiments, this study aimed to understand the influence of architectural variations on flap outcome. METHODS: Fasciocutaneous flaps were designed on the ears of New Zealand rabbits, and the animals were randomized into four groups having flaps that used the larger anterior marginal vein (1.3 mm) or the smaller central vein (0.6 mm) for arterial inflow, with or without isolation of the flap from its bed with a silicone sheet. Flaps were observed for area of flap survival and vasculature was assessed by microangiography. RESULTS: Using the smaller central vein for arterial inflow (n = 15), arterialized venous flaps had an excellent outcome, with good flap survival in 100 percent of the animals (survival of >85 percent of flap area), and a mean flap survival area of 99.4 +/- 1.6 percent. Even when neovascularization was prevented by isolation of the flaps (n = 14), 92 percent of central vein flaps showed good survival, with a mean flap survival area of 93.3 +/- 7.3 percent, which was significantly better than that of anterior marginal vein flaps (n = 22), which showed good flap survival in only 27 percent of the animals (mean flap survival area, 76.4 +/- 12.1 percent). CONCLUSIONS: Survival of arterialized venous flaps is optimized by using smaller-caliber veins for inflow and reserving larger-caliber veins for outflow. This regulates inflow and avoids high blood pressure, and arterialized venous flaps behave as physiologic flaps do, by not relying on neovascularization for survival.

Mixed arterio-venous insufficiency in random skin flaps in the rat: is the application of medicinal leeches beneficial?

BACKGROUND: Leeches are commonly used in reconstructive surgery for the treatment of venous congestion in flaps. Documented mechanisms of action are (1) injection of the anticoagulant hirudin; (2) active suction of blood; and (3) passive oozing of the bite wound. Even though the benefits of leeches in venous congestion are widely accepted, little is known about their effects in mixed arterio-venous insufficiency. METHODS: Thirty Wistar rats were randomized into three groups (n = 10 each), and 9 x 3 cm ischemic random skin flaps were elevated on the rat dorsum. Group 1 served as controls. In group 2, one leech was applied to the distal part of the flap and in group 3, three leeches were applied, one at a time, at 8 h intervals. Postoperatively, flap survival and perfusion were quantitated by daily planimetry and laser-Doppler imaging. RESULTS: The application of a single leech on postoperative day 7 did not lead to any statistically significant changes in total flap survival or tissue perfusion. Triple leech application, however, caused a significant decrease in flap survival of 6.6% at day 7 (3x leech versus control: 45.8% +/- 8.5% versus 52.4% +/- 8.5%, respectively) (P < 0.01) and a decline in flap perfusion of 13% (3x leech versus control: 71.3% +/- 16.6% versus 84.3% +/- 9.3%, respectively) (P < 0.01). CONCLUSION: The data demonstrate that the application of leeches in the setting of mixed arterio-venous insufficiency can be hazardous to flap viability. Pure venous congestion with an adequate arterial supply remains the only indication for controlled leech application in the clinical setting.

Progressive tissue injury in burns is reduced by rNAPc2.

INTRODUCTION: Burn wounds are characterised by central necrosis surrounded by an area of stasis with compromised perfusion. Secondary aggravation of the burn wound due to ischaemia in the zone of stasis can also result in necrosis. This study aims to improve circulation in the zone of stasis by reducing microthrombus formation and thereby to reduce secondary aggravation. MATERIAL AND METHODS: Recombinant nematode anticoagulant protein (rNAPc2) was administered to Wistar rats at 3 or 30 microg/kg as a single or daily dose. A comb pattern burn was induced on the dorsum of these rats and its evolution monitored by serial photography, planimetry, laser doppler flowmetry and immunohistochemistry. RESULTS: In the 30 microg/kg daily group, extension of the burn wound was curbed, limiting the burn area to 1.99+/-0.67 cm(2) on day 28, compared to 3.51+/-0.37 cm(2) in the control group (p=0.015). Laser doppler evaluation showed a significant (p<0.001) increase in circulation in the first day post-burn. Significantly less (p<0.001) microvascular fibrin formation was observed by immunohistochemistry. CONCLUSION: Anticoagulation with rNAPc2 improved perfusion of the burn wound. The resultant reduction in the area of the burn led to earlier healing and less scar contracture.

The free serratus anterior flap and its cutaneous component for reconstruction of the face: a series of 27 cases.

BACKGROUND: The serratus anterior flap is commonly used without its cutaneous component and is covered with a skin graft. The authors have successfully used the free serratus anterior flap along with its skin paddle and have found it to be valuable for reconstruction of the face. METHODS: Fresh cadaveric dissections and arteriography were performed to identify perforator vessels to the skin overlying the muscle. Clinically, free transfer of the musculocutaneous flap to the face was carried out in 27 patients, mostly for severe noma (infection) sequelae. RESULTS: Anatomical dissection and arteriography revealed no cutaneous perforator vessels directly communicating with the vascular pedicle of the muscle. However, large perforators from the intercostal vessels were found passing through the muscle to reach the skin. In the clinical cases, flap survival was 100 percent in 24 patients. CONCLUSIONS: The serratus anterior musculocutaneous flap is reliable and particularly well-suited for reconstruction of the face, and has many advantages. The authors speculate that the skin paddle may be vascularized by perforators from the intercostal vessels communicating with the thoracodorsal pedicle through intramuscular choke vessels.

Active-site inactivated FVIIa decreases thrombosis and necrosis in a random skin flap model of acute ischemia.

BACKGROUND: Previous studies have emphasized the role of ischemia in inducing vascular thrombosis. MATERIALS AND METHODS: Using a skin flap model of acute ischemia in the rat, we studied the effect of active-site inactivated factor VIIa (FVIIai), an inhibitor of tissue factor (TF), on tissue survival during acute ischemia. RESULTS: Ribonuclease protection analysis revealed an increase in TF in ischemic parts of the flap, and in situ hybridization localized this increase mainly to perivascular cells. A decrease in vascular thrombosis, as determined by fibrin immunostaining, was observed in FVIIai-treated animals. Intravenous administration of FVIIai had a positive impact on survival of the flap. Laser Doppler flowmetry revealed an increase in blood flow in the FVIIai-treated group. In treated animals, prothrombin time (PT) was increased (P < 0.01), whereas partial thromboplastin time (APTT) was unaltered; no significant impairment in systemic hemostasis (peri- and postoperative bleeding) was observed. CONCLUSIONS: These findings demonstrate that TF expression is increased in perivascular cells in ischemic skin flaps and that FVIIai, by inhibiting TF, increases flap survival.