Nontopical Nitrates in Flap Perfusion and Delay Phenomenon.

Background: Topical nitrates have demonstrated efficacy in improving flap perfusion. However, evidence for nontopical nitrates in modulation of flap perfusion dynamics has yet to be consolidated. Here, we review evidence regarding the use of intravascular, sublingual, and oral nitrates in modulating flap perfusion. Methods: We performed a review of the literature for evidence linking nontopical nitrates and flap perfusion, and included clinical studies, animal studies, and in vitro studies. Results: Evidence suggests that intravascular, sublingual, and oral nitrates exert vasodilatory properties, which may be harnessed for identification of perforators and improved flap perfusion. We also found evidence suggesting nitrates may facilitate ischemic preconditioning while reducing ischemia-reperfusion injury. Conclusions: Nitrates delivered intravascularly, sublingually, or orally may increase flap perfusion and serve as a method for ischemic preconditioning, particularly in the intraoperative setting.

Analysis of Major and Minor Pedicles in Flap Perfusion by Computational Fluid Dynamics.

Background: Type 2 muscle flaps are characterized by major and minor pedicles, such that the minor pedicle is unreliable, and the major pedicle is a requirement for the success of the flap. The role of the minor pedicle, beyond the decreased caliber and decreased vascular territory in comparison to the major pedicle, is poorly understood. We sought to model the fluid dynamics of a model flap containing a major and minor pedicle to understand differences between the pedicles and the implications on perfusion. Methods: We first generated a computer-assisted design model of a type 2 flap with a major and minor pedicle. We then performed computational fluid dynamics to analyze velocities and flow within the pedicles and flap. Results: In our investigation, we found that the flow velocity within the major pedicle was higher than the minor pedicle, indicative of decreased resistance to flow. Concomitantly, we found decreased pressures within the major pedicle, reflecting decreasing resistance to flow. Interestingly, we found increased kinematic viscosity in flap areas supplied by the minor pedicle, suggesting decreased flow rates and increased resistance. Conclusions: We identified that the major pedicle has increased flow velocity, decreased resistance, and decreased kinematic viscosity, suggesting its dominance in maintaining flap perfusion. Our study also identifies computational fluid dynamics as a powerful tool in studying flap perfusion dynamics.

Modulation of Cellular Stemness for Enhanced Fat Grafting.

Low volumetric retention limits the utility of fat grafting. Although inclusion of stem cells and platelet-rich plasma have been proposed to enhance graft retention, accumulating evidence has failed to show a clear benefit. Here, we propose a strategy to pharmacologically enhance stemness of stem and progenitor cell populations in fat grafts to promote increased volume retention and tissue health. We also propose how to integrate stemness-promoting and differentiation-promoting therapies such as platelet-rich plasma, and viability promoting therapies within the common fat grafting workflow to achieve optimal fat grafting results.

Management of partial-thickness rotator cuff tears with autologous adipose-derived regenerative cells is safe and more effective than injection of corticosteroid.

Symptomatic, partial-thickness rotator cuff tears (sPTRCT) are problematic. This study tested the hypothesis that management of sPTRCT with injection of fresh, uncultured, unmodified, autologous, adipose-derived regenerative cells (UA-ADRCs) is safe and more effective than injection of corticosteroid even in the long run. To this end, subjects who had completed a former randomized controlled trial were enrolled in the present study. At baseline these subjects had not responded to physical therapy treatments for at least 6 weeks, and were randomly assigned to receive respectively a single injection of UA-ADRCs (n = 11) or a single injection of methylprednisolone (n = 5). Efficacy was assessed using the ASES Total score, pain visual analogue scale (VAS), RAND Short Form-36 Health Survey and range of motion at 33.2 ± 1.0 (mean ± SD) and 40.6 ± 1.9 months post-treatment. Proton density, fat-saturated, T2-weighted MRI of the index shoulder was performed at both study visits. There were no greater risks connected with injection of UA-ADRCs than those connected with injection of corticosteroid. The subjects in the UA-ADRCs group showed statistically significantly higher mean ASES Total scores than the subjects in the corticosteroid group. The MRI scans at 6 months post-treatment allowed to "watch the UA-ADRCs at work".

Unfolded protein response and angiogenesis in malignancies.

Cellular stress, arising from accumulation of unfolded proteins, occurs frequently in rapidly proliferating cancer cells. This cellular stress, in turn, activates the unfolded protein response (UPR), an interconnected set of signal transduction pathways that alleviate the proteostatic stress. The UPR is implicated in cancer cell survival and proliferation through upregulation of pro-tumorigenic pathways that ultimately promote malignant metabolism and neoangiogenesis. Here, we reviewed mechanisms of signaling crosstalk between the UPR and angiogenesis pathways, as well as transmissible ER stress and the role in tumor growth and development. To characterize differences in UPR and UPR-mediated angiogenesis in malignancy, we employed a data mining approach using patient tumor data from The Cancer Genome Atlas (TCGA). The analysis of TCGA revealed differences in UPR between malignant samples versus their non-malignant counterparts.

Targeting TRAF3IP2 inhibits angiogenesis in glioblastoma.

Increased vascularization, also known as neoangiogenesis, plays a major role in many cancers, including glioblastoma multiforme (GBM), by contributing to their aggressive growth and metastasis. Although anti-angiogenic therapies provide some clinical improvement, they fail to significantly improve the overall survival of GBM patients. Since various pro-angiogenic mediators drive GBM, we hypothesized that identifying targetable genes that broadly inhibit multiple pro-angiogenic mediators will significantly promote favorable outcomes. Here, we identified TRAF3IP2 (TRAF3-interacting protein 2) as a critical regulator of angiogenesis in GBM. We demonstrated that knockdown of TRAF3IP2 in an intracranial model of GBM significantly reduces vascularization. Targeting TRAF3IP2 significantly downregulated VEGF, IL6, ANGPT2, IL8, FZGF2, PGF, IL1ß, EGF, PDGFRB, and VEGFR2 expression in residual tumors. Our data also indicate that exogenous addition of VEGF partially restores angiogenesis by TRAF3IP2-silenced cells, suggesting that TRAF3IP2 promotes angiogenesis through VEGF- and non-VEGF-dependent mechanisms. These results indicate the anti-angiogenic and anti-tumorigenic potential of targeting TRAF3IP2 in GBM, a deadly cancer with limited treatment options.

Biallelic, Selectable, Knock-in Targeting of CCR5 via CRISPR-Cas9 Mediated Homology Directed Repair Inhibits HIV-1 Replication.

Transplanting HIV-1 positive patients with hematopoietic stem cells homozygous for a 32 bp deletion in the chemokine receptor type 5 (CCR5) gene resulted in a loss of detectable HIV-1, suggesting genetically disrupting CCR5 is a promising approach for HIV-1 cure. Targeting the CCR5-locus with CRISPR-Cas9 was shown to decrease the amount of CCR5 expression and HIV-1 susceptibility in vitro as well as in vivo. Still, only the individuals homozygous for the CCR5-Δ32 frameshift mutation confer complete resistance to HIV-1 infection. In this study we introduce a mechanism to target CCR5 and efficiently select for cells with biallelic frameshift insertion, using CRISPR-Cas9 mediated homology directed repair (HDR). We hypothesized that cells harboring two different selectable markers (double positive), each in one allele of the CCR5 locus, would carry a frameshift mutation in both alleles, lack CCR5 expression and resist HIV-1 infection. Inducing double-stranded breaks (DSB) via CRISPR-Cas9 leads to HDR and integration of a donor plasmid. Double-positive cells were selected via fluorescence-activated cell sorting (FACS), and CCR5 was analyzed genetically, phenotypically, and functionally. Targeted and selected populations showed a very high frequency of mutations and a drastic reduction in CCR5 surface expression. Most importantly, double-positive cells displayed potent inhibition to HIV-1 infection. Taken together, we show that targeting cells via CRISPR-Cas9 mediated HDR enables efficient selection of mutant cells that are deficient for CCR5 and highly resistant to HIV-1 infection.

Why and how to use the body's own stem cells for regeneration in musculoskeletal disorders: a primer.

BACKGROUND: Recently, the management of musculoskeletal disorders with the patients' own stem cells, isolated from the walls of small blood vessels, which can be found in great numbers in the adipose tissue, has received considerable attention. On the other hand, there are still misconceptions about these adipose-derived regenerative cells (ADRCs) that contain vascular-associated pluripotent stem cells (vaPS cells) in regenerative medicine. METHODS: Based on our previous publications on this topic, we have developed a concept to describe the significance of the ADRCs/vaPS cells in the field of orthobiologics as briefly as possible and at the same time as precisely as possible. RESULTS: The ADRCs/vaPS cells belong to the group of orthobiologics that are based on autologous cells. Because the latter can both stimulate a patient's body's localized self-healing power and provide new cells that can integrate into the host tissue during the healing response when the localized self-healing power is exhausted, this group of orthobiologics appears more advantageous than cell-free orthobiologics and orthobiologics that are based on allogeneic cells. Within the group of orthobiologics that are based on autologous cells, enzymatically isolated, uncultured ADRCs/vaPS cells have several advantages over non-enzymatically isolated cells/microfragmented fat as well as over uncultured bone marrow aspirate concentrate and cultured cells (adipose-derived stem cells, bone marrow-derived mesenchymal stem cells). CONCLUSIONS: The use of ADRCs/vaPS cells can be seamlessly integrated into modern orthopedic treatment concepts, which can be understood as the optimization of a process which-albeit less efficiently-also takes place physiologically. Accordingly, this new safe and effective type of treatment is attractive in terms of holistic thinking and personalized medicine.

First immunohistochemical evidence of human tendon repair following stem cell injection: A case report and review of literature.

BACKGROUND: Current clinical treatment options for symptomatic, partial-thickness rotator cuff tear (sPTRCT) offer only limited potential for true tissue healing and improvement of clinical results. In animal models, injections of adult stem cells isolated from adipose tissue into tendon injuries evidenced histological regeneration of tendon tissue. However, it is unclear whether such beneficial effects could also be observed in a human tendon treated with fresh, uncultured, autologous, adipose derived regenerative cells (UA-ADRCs). A specific challenge in this regard is that UA-ADRCs cannot be labeled and, thus, not unequivocally identified in the host tissue. Therefore, histological regeneration of injured human tendons after injection of UA-ADRCs must be assessed using comprehensive, immunohistochemical and microscopic analysis of biopsies taken from the treated tendon a few weeks after injection of UA-ADRCs. CASE SUMMARY: A 66-year-old patient suffered from sPTRCT affecting the right supraspinatus and infraspinatus tendon, caused by a bicycle accident. On day 18 post injury [day 16 post magnetic resonance imaging (MRI) examination] approximately 100 g of abdominal adipose tissue was harvested by liposuction, from which approximately 75 × 106 UA-ADRCs were isolated within 2 h. Then, UA-ADRCs were injected (controlled by biplanar X-ray imaging) adjacent to the injured supraspinatus tendon immediately after isolation. Despite fast clinical recovery, a follow-up MRI examination 2.5 mo post treatment indicated the need for open revision of the injured infraspinatus tendon, which had not been treated with UA-ADRCs. During this operation, a biopsy was taken from the supraspinatus tendon at the position of the injury. A comprehensive, immunohistochemical and microscopic analysis of the biopsy (comprising 13 antibodies) was indicative of newly formed tendon tissue. CONCLUSION: Injection of UA-ADRCs can result in regeneration of injured human tendons by formation of new tendon tissue.

Increased Efficiency for Biallelic Mutations of the CCR5 Gene by CRISPR-Cas9 Using Multiple Guide RNAs As a Novel Therapeutic Option for Human Immunodeficiency Virus.

CCR5 is a coreceptor of human immunodeficiency virus type 1 (HIV-1). Transplantation of hematopoietic stem cells homozygous for a 32-bp deletion in CCR5 resulted in a loss of detectable HIV-1 in two patients, suggesting that genetic strategies to knockout CCR5 expression would be a promising gene therapy approach for HIV-1-infected patients. In this study, we targeted CCR5 by CRISPR-Cas9 with a single-guide (sgRNA) and observed 35% indel frequency. When we expressed hCas9 and two gRNAs, the Surveyor assay showed that Cas9-mediated cleavage was increased by 10% with two sgRNAs. Genotype analysis on individual clones showed 11 of 13 carried biallelic mutations, where 4 clones had frameshift (FS) mutations. Taken together, these results indicate that the efficiency of biallelic FS mutations and the knockout of the CCR5 necessary to prevent viral replication were significantly increased with two sgRNAs. These studies demonstrate the knockout of CCR5 and the potential for translational development.

Identification and Targeting of Thomsen-Friedenreich and IL1RAP Antigens on Chronic Myeloid Leukemia Stem Cells Using Bi-Specific Antibodies.

INTRODUCTION: Quiescent leukemia stem cells (LSCs) play a major role in therapeutic resistance and disease progression of chronic myeloid leukemia (CML). LSCs belong to the primitive population; CD34+CD38-Lin-, which does not distinguish normal hematopoietic stem cells (HSC) from CML LSCs. Because Thomsen-Friedenreich/CD176 antigen is expressed on CD34+ HSC and IL1RAP is tightly correlated to BCR-ABL expression, we sought to increase the specificity towards LSC by using additional biomarkers. METHODS: We evaluated the co-expression of both antigens on CD34+ peripheral blood mononuclear cells (PBMCs) from both healthy volunteers and CML patients, using flow cytometry. Then, we used site-directed mutagenesis to induce knob-in-hole mutations in the human IgG heavy chain and the human lambda light chain to generate the bi-specific antibody (Bis-Ab) TF/RAP that binds both antigens simultaneously. We measured complement-directed cytotoxicity (CDC) in CML samples with the Bis-Ab by flow cytometry. RESULTS: In contrast to healthy volunteers, CML samples displayed a highly significant co-expression of CD176 and IL1RAP. When either a double-positive cell line or CML samples were treated with increasing doses of Bis-Ab, increased binding and CDC was observed indicating co-operative binding of the Bis-Ab as compared to monoclonal antibodies. DISCUSSION: These results show that the bi-specific antibody is capable of targeting IL1RAP+ and CD176+ cell population among CML PBMCs, but not corresponding normal cells in CDC assay. We hereby offer a novel strategy for the depletion of CML stem cells from the bulk population in clinical hematopoietic stem cell transplantation.

Anti-Cancer and Anti-Bacterial Effects of Terfezia boudieri-Derived Silver Nanoparticles.

BACKGROUND: The use of nanoparticles has markedly increased in biomedical sciences. The silver nanoparticles (AgNPs) have been investigated for their applicability to deliver chemotherapeutic/antibacterial agents to treat cancer or infections disease. However, the existing chemical and physical methods of synthesizing AgNPs are considered inefficient, expensive and toxic. METHODS: Natural products have emerged as viable candidates for nanoparticle production, including the use of Terfezia boudieri (T. boudieri), a member of the edible truffle family. Accordingly, our goal was to synthesize AgNPs using an aqueous extract of T. boudieri (green synthesized AgNPs). Since certain infectious agents are linked to cancer, we investigated their potential as anti-cancer and antibacterial agents. RESULTS: The synthesis of AgNPs was confirmed by the presence of an absorption peak at 450nm by spectroscopy. The physico-chemical properties of green synthesized AgNPs were analyzed by UV-Vis, FT-IR, XRD, SEM, and TEM. In addition, their potential to inhibit cancer cell (proliferation and the growth of infectious bacteria were investigated. CONCLUSION: The size of nanoparticles ranged between 20-30nm. They exerted significant cytotoxicity and bactericidal effects in a concentration and time-dependent manner compared to T. boudieri extract alone. Interestingly, the synthesis of smaller AgNPs was correlated with longer synthesis time and enhanced cytotoxic and bactericidal properties.

Targeting TRAF3IP2, Compared to Rab27, is More Effective in Suppressing the Development and Metastasis of Breast Cancer.

Here we investigated the roles of Rab27a, a player in exosome release, and TRAF3IP2, an inflammatory mediator, in development and metastasis of breast cancer (BC) in vivo. Knockdown (KD) of Rab27a (MDAKDRab27a) or TRAF3IP2 (MDAKDTRAF3IP2) in triple negative MDA-MB231 cells reduced tumor growth by 70-97% compared to wild-type tumors (MDAw). While metastasis was detected in MDAw-injected animals, none was detected in MDAKDRab27a- or MDAKDTRAF3IP2-injected animals. Interestingly, micrometastasis was detected only in the MDAKDRab27a-injected group. In addition to inhibiting tumor growth and metastasis, silencing TRAF3IP2 disrupted inter-cellular inflammatory mediator-mediated communication with mesenchymal stem cells (MSCs) injected into contralateral mammary gland, evidenced by the lack of tumor growth at MSC-injected site. Of translational significance, treatment of pre-formed MDAw-tumors with a lentiviral-TRAF3IP2-shRNA not only regressed their size, but also prevented metastasis. These results demonstrate that while silencing Rab27a and TRAF3IP2 each inhibited tumor growth and metastasis, silencing TRAF3IP2 is more effective; targeting TRAF3IP2 inhibited tumor formation, regressed preformed tumors, and prevented both macro- and micrometastasis. Silencing TRAF3IP2 also blocked interaction between tumor cells and MSCs injected into the contralateral gland, as evidenced by the lack of tumor formation on MSCs injected site. These results identify TRAF3IP2 as a novel therapeutic target in BC.

Towards a Comprehensive Understanding of UA-ADRCs (Uncultured, Autologous, Fresh, Unmodified, Adipose Derived Regenerative Cells, Isolated at Point of Care) in Regenerative Medicine.

It has become practically impossible to survey the literature on cells derived from adipose tissue for regenerative medicine. The aim of this paper is to provide a comprehensive and translational understanding of the potential of UA-ADRCs (uncultured, unmodified, fresh, autologous adipose derived regenerative cells isolated at the point of care) and its application in regenerative medicine. We provide profound basic and clinical evidence demonstrating that tissue regeneration with UA-ADRCs is safe and effective. ADRCs are neither 'fat stem cells' nor could they exclusively be isolated from adipose tissue. ADRCs contain the same adult stem cells ubiquitously present in the walls of blood vessels that are able to differentiate into cells of all three germ layers. Of note, the specific isolation procedure used has a significant impact on the number and viability of cells and hence on safety and efficacy of UA-ADRCs. Furthermore, there is no need to specifically isolate and separate stem cells from the initial mixture of progenitor and stem cells found in ADRCs. Most importantly, UA-ADRCs have the physiological capacity to adequately regenerate tissue without need for more than minimally manipulating, stimulating and/or (genetically) reprogramming the cells for a broad range of clinical applications. Tissue regeneration with UA-ADRCs fulfills the criteria of homologous use as defined by the regulatory authorities.

Safety and efficacy of treating symptomatic, partial-thickness rotator cuff tears with fresh, uncultured, unmodified, autologous adipose-derived regenerative cells (UA-ADRCs) isolated at the point of care: a prospective, randomized, controlled first-in-human pilot study.

BACKGROUND: This study tested the hypothesis that treatment of symptomatic, partial-thickness rotator cuff tears (sPTRCT) with fresh, uncultured, unmodified, autologous adipose-derived regenerative cells (UA-ADRCs) isolated from lipoaspirate at the point of care is safe and more effective than corticosteroid injection. METHODS: Subjects aged between 30 and 75 years with sPTRCT who did not respond to physical therapy treatments for at least 6 weeks were randomly assigned to receive a single injection of an average 11.4 × 106 UA-ADRCs (in 5 mL liquid; mean cell viability: 88%) (n = 11; modified intention-to-treat (mITT) population) or a single injection of 80 mg of methylprednisolone (40 mg/mL; 2 mL) plus 3 mL of 0.25% bupivacaine (n = 5; mITT population), respectively. Safety and efficacy were assessed using the American Shoulder and Elbow Surgeons Standardized Shoulder Assessment Form (ASES), RAND Short Form-36 Health Survey, and pain visual analogue scale (VAS) at baseline (BL) as well as 3 weeks (W3), W6, W9, W12, W24, W32, W40, and W52 post treatment. Fat-saturated T2-weighted magnetic resonance imaging of the shoulder was performed at BL as well as at W24 and W52 post treatment. RESULTS: No severe adverse events related to the injection of UA-ADRCs were observed in the 12 months post treatment. The risks connected with treatment of sPTRCT with UA-ADRCs were not greater than those connected with treatment of sPTRCT with corticosteroid injection. However, one subject in the corticosteroid group developed a full rotator cuff tear during the course of this pilot study. Despite the small number of subjects in this pilot study, those in the UA-ADRCs group showed statistically significantly higher mean ASES total scores at W24 and W52 post treatment than those in the corticosteroid group (p < 0.05). DISCUSSION: This pilot study suggests that the use of UA-ADRCs in subjects with sPTRCT is safe and leads to improved shoulder function without adverse effects. To verify the results of this initial safety and feasibility pilot study in a larger patient population, a randomized controlled trial on 246 patients suffering from sPTRCT is currently ongoing. TRIAL REGISTRATION: Clinicaltrials.gov ID NCT02918136. Registered September 28, 2016, https://clinicaltrials.gov/ct2/show/NCT02918136. LEVEL OF EVIDENCE: Level I; prospective, randomized, controlled trial.

A New Humanized Mouse Model Mimics Humans in Lacking α-Gal Epitopes and Secreting Anti-Gal Antibodies.

Mice have been used as accepted tools for investigating complex human diseases and new drug therapies because of their shared genetics and anatomical characteristics with humans. However, the tissues in mice are different from humans in that human cells have a natural mutation in the α1,3 galactosyltransferase (α1,3GT) gene and lack α-Gal epitopes on glycosylated proteins, whereas mice and other nonprimate mammals express this epitope. The lack of α-Gal epitopes in humans results in the loss of immune tolerance to this epitope and production of abundant natural anti-Gal Abs. These natural anti-Gal Abs can be used as an adjuvant to enhance processing of vaccine epitopes to APCs. However, wild-type mice and all existing humanized mouse models cannot be used to test the efficacy of vaccines expressing α-Gal epitopes because they express α-Gal epitopes and lack anti-Gal Abs. Therefore, in an effort to bridge the gap between the mouse models and humans, we developed a new humanized mouse model that mimics humans in that it lacks α-Gal epitopes and secretes human anti-Gal Abs. The new humanized mouse model (Hu-NSG/α-Galnull) is designed to be used for preclinical evaluations of viral and tumor vaccines based on α-Gal epitopes, human-specific immune responses, xenotransplantation studies, and in vivo biomaterials evaluation. To our knowledge, our new Hu-NSG/α-Galnull is the first available humanized mouse model with such features.

Isolation of adipose tissue derived regenerative cells from human subcutaneous tissue with or without the use of an enzymatic reagent.

Freshly isolated, uncultured, autologous adipose derived regenerative cells (ADRCs) have emerged as a promising tool for regenerative cell therapy. The Transpose RT system (InGeneron, Inc., Houston, TX, USA) is a system for isolating ADRCs from adipose tissue, commercially available in Europe as a CE-marked medical device and under clinical evaluation in the United States. This system makes use of the proprietary, enzymatic Matrase Reagent for isolating cells. The present study addressed the question whether the use of Matrase Reagent influences cell yield, cell viability, live cell yield, biological characteristics, physiological functions or structural properties of the ADRCs in final cell suspension. Identical samples of subcutaneous adipose tissue from 12 subjects undergoing elective lipoplasty were processed either with or without the use of Matrase Reagent. Then, characteristics of the ADRCs in the respective final cell suspensions were evaluated. Compared to non-enzymatic isolation, enzymatic isolation resulted in approximately twelve times higher mean cell yield (i.e., numbers of viable cells/ml lipoaspirate) and approximately 16 times more colony forming units. Despite these differences, cells isolated from lipoaspirate both with and without the use of Matrase Reagent were independently able to differentiate into cells of all three germ layers. This indicates that biological characteristics, physiological functions or structural properties relevant for the intended use were not altered or induced using Matrase Reagent. A comprehensive literature review demonstrated that isolation of ADRCs from lipoaspirate using the Transpose RT system and the Matrase Reagent results in the highest viable cell yield among published data regarding isolation of ADRCs from lipoaspirate.

Improved guided bone regeneration by combined application of unmodified, fresh autologous adipose derived regenerative cells and plasma rich in growth factors: A first-in-human case report and literature review.

BACKGROUND: Novel strategies are needed for improving guided bone regeneration (GBR) in oral surgery prior to implant placement, particularly in maxillary sinus augmentation (GBR-MSA) and in lateral alveolar ridge augmentation (LRA). This study tested the hypothesis that the combination of freshly isolated, unmodified autologous adipose-derived regenerative cells (UA-ADRCs), fraction 2 of plasma rich in growth factors (PRGF-2) and an osteoinductive scaffold (OIS) (UA-ADRC/PRGF-2/OIS) is superior to the combination of PRGF-2 and the same OIS alone (PRGF-2/OIS) in GBR-MSA/LRA. CASE SUMMARY: A 79-year-old patient was treated with a bilateral external sinus lift procedure as well as a bilateral lateral alveolar ridge augmentation. GBR-MSA/LRA was performed with UA-ADRC/PRGF-2/OIS on the right side, and with PRGF-2/OIS on the left side. Biopsies were collected at 6 wk and 34 wk after GBR-MSA/LRA. At the latter time point implants were placed. Radiographs (32 mo follow-up time) demonstrated excellent bone healing. No radiological or histological signs of inflammation were observed. Detailed histologic, histomorphometric, and immunohistochemical analysis of the biopsies evidenced that UA-ADRC/PRGF-2/OIS resulted in better and faster bone regeneration than PRGF-2/OIS. CONCLUSION: GBR-MSA with UA-ADRCs, PRGF-2, and an OIS shows effectiveness without adverse effects.

Breast Tumor Microenvironment Can Transform Naive Mesenchymal Stem Cells into Tumor-Forming Cells in Nude Mice.

How mesenchymal stem cells (MSCs) interact with tumor cells and promote tumor growth is not well understood. In this study, we demonstrate that when naive MSCs and malignant breast cancer cells (MDA-MB231) were injected into opposing mammary glands of an immunodeficient nude mouse, both cell types formed tumor-like masses within 8 weeks at the injected site. Surprisingly, MDA-MB231 cells were detected in the opposing mammary gland injected with the naive MSCs, indicating migration and crosstalk between naive MSCs and MDA-MB231 cells. Furthermore, when naive MSCs preexposed to MDA-MB231-derived conditioned medium (CM; MSCCM) or purified exosomes (Exo; MSCExo) were injected into mammary glands of nude mice, they too formed a tumor-like mass with stromal tissue within 14 weeks. Interestingly, cells dissociated from these primary explants also formed tumor-like masses. Finally, injecting MSCCM or MSCExo and naive MSCs into opposing mammary glands formed tumor-like masses on the naive MSC-injected side, suggesting migration and crosstalk between MSCCM or MSCExo with naive MSCs, similar to that observed between malignant MDA-MB231 cells and naive MSCs. Importantly, molecular analysis of MSCCM and MSCExo revealed DNA hypermethylation. These data demonstrate that MSCs and breast cancer cells communicate, resulting in the transformation of naive MSCs into cells capable of forming explants in nude mice. Our data also suggest that DNA hypermethylation might have contribute to their migration. Understanding the crosstalk between MSCs and tumor cells, and identifying the players involved in their interaction, will help us develop novel therapeutics for breast cancer regression and elimination.