No Clinical Efficacy of Adipose-Derived Regenerative Cells and Lipotransfer in Breast Cancer-Related Lymphedema: A double-blinded placebo-controlled phase-II trial.

BACKGROUND: Breast cancer-related lymphedema (BCRL) is a debilitating sequela affecting up to one in three breast cancer survivors. Current treatments are palliative and does not address the underlying lymphatic injury. Recently, preclinical and non-randomized studies have shown promising results using Adipose-Derived Regenerative Cells (ADRCs) and lipotransfer in alleviating BCRL through regeneration of lymphatic tissue. However no randomized controlled trial has been performed in an attempt to eliminate a placebo effect. METHODS: This randomized, double-blinded, placebo-controlled trial included patients with no-option, persistent disabling unilateral BCRL. Patients were randomly assigned in a 1:1 ratio to receive either autologous ADRCs (4.20x10 7±1.75x10 7 cells) and 30cc lipotransfer or placebo (saline) to the axilla. The primary outcome was a change in BCRL volume one year after treatment. Secondary outcomes included changes in the quality of life, indocyanine green lymphangiography stage, bioimpedance, and safety. RESULTS: Eighty patients were included, of which 39 were allocated to ADRCs and lipotransfer treatment and 41 to placebo treatment. Baseline characteristics were similar in both groups. One year after treatment, no objective improvements were observed in the treatment or placebo groups. In contrast, significant subjective improvements were noted for both the treatment and placebo groups. CONCLUSION: This trial failed to confirm a benefit of ADRCs and lipotransfer in the treatment of BCRL. These non-confirmatory results suggest that ADRC and lipotransfer should not be recommended for alleviating BCRL at this time. However, we cannot exclude that repeated treatments or higher doses of ADRCs or lipotransfer could yield a clinical effect.

CD31 defines a subpopulation of human adipose-derived regenerative cells with potent angiogenic effects.

Cellular heterogeneity represents a major challenge for regenerative treatment using freshly isolated Adipose Derived Regenerative Cells (ADRCs). Emerging data suggest superior efficacy of ADRCs as compared to the ex vivo expanded and more homogeneous ADRCs (= ASCs) for indications involving (micro)vascular deficiency, however, it remains unknown which ADRC cell subtypes account for the improvement. Surprisingly, we found regarding erectile dysfunction (ED) that the number of injected CD31+ ADRCs correlated positively with erectile function 12 months after one bolus of autologous ADRCs. Comprehensive in vitro and ex vivo analyses confirmed superior pro-angiogenic and paracrine effects of human CD31+ enriched ADRCs compared to the corresponding CD31- and parent ADRCs. When CD31+, CD31- and ADRCs were co-cultured in aortic ring- and corpus cavernous tube formation assays, the CD31+ ADRCs induced significantly higher tube development. This effect was corroborated using conditioned medium (CM), while quantitative mass spectrometric analysis suggested that this is likely explained by secretory pro-angiogenic proteins including DKK3, ANGPT2, ANAX2 and VIM, all enriched in CD31+ ADRC CM. Single-cell RNA sequencing showed that transcripts of the upregulated and secreted proteins were present in 9 endothelial ADRC subsets including endothelial progenitor cells in the heterogenous non-cultured ADRCs. Our data suggest that the vascular benefit of using ADRCs in regenerative medicine is dictated by CD31+ ADRCs.

Prevalence of prostate cancer in men with haematuria: a systematic review and meta-analysis.

OBJECTIVES: To investigate the prevalence of prostate cancer in men attending evaluation for haematuria, as this could help healthcare providers to determine whether men with haematuria should have prostate examinations performed. METHODS: The study was performed according to a pre-specified protocol uploaded to the International Prospective Register of Systematic Reviews (PROSPERO; CRD42022299383). A systematic search of MEDLINE, Ovid and Google Scholar was performed in December 2021. Two independent researchers evaluated all titles, available abstracts, and full texts. We included studies on adult men (aged ≥18 years) describing haematuria and prostate cancer. RESULTS: We screened 4252 titles and abstracts when available and assessed 350 studies in full text. In total, 65 studies were included and 42 was summarised in a meta-analysis. In total, 18 752 men with haematuria were included, and the pooled prevalence (95% confidence interval [CI]) of prostate cancer was 3.0% (2.0-4.1%). In men with macroscopic haematuria, the pooled prevalence (95% CI) of prostate cancer was 5.9% (2.9-9.9%; n = 265/5373). In men with microscopic haematuria, the pooled prevalence (95% CI) of prostate cancer was 1.4% (0.8-2.2%; n = 71/6642). CONCLUSION: Our findings indicate that the prevalence of prostate cancer is considerable in men attending evaluation for haematuria. Therefore, digital rectal examination and prostate-specific antigen measurement should become a standard procedure for all men with haematuria, especially for men with macroscopic haematuria.

Adipose-Derived Stem Cells from Type 2 Diabetic Rats Retain Positive Effects in a Rat Model of Erectile Dysfunction.

Erectile dysfunction is a common complication associated with type 2 diabetes mellitus (T2DM) and after prostatectomy in relation to cancer. The regenerative effect of cultured adipose-derived stem cells (ASCs) for ED therapy has been documented in multiple preclinical trials as well as in recent Pase 1 trials in humans. However, some studies indicate that diabetes negatively affects the mesenchymal stem cell pool, implying that ASCs from T2DM patients could have impaired regenerative capacity. Here, we directly compared ASCs from age-matched diabetic Goto-Kakizaki (ASCGK) and non-diabetic wild type rats (ASCWT) with regard to their phenotypes, proteomes and ability to rescue ED in normal rats. Despite ASCGK exhibiting a slightly lower proliferation rate, ASCGK and ASCWT proteomes were more or less identical, and after injections to corpus cavernosum they were equally efficient in restoring erectile function in a rat ED model entailing bilateral nerve crush injury. Moreover, molecular analysis of the corpus cavernosum tissue revealed that both ASCGK and ASCWT treated rats had increased induction of genes involved in recovering endothelial function. Thus, our finding argues that T2DM does not appear to be a limiting factor for autologous adipose stem cell therapy when correcting for ED.

Small diameter polycaprolactone vascular grafts are patent in sheep carotid bypass but require antithrombotic therapy.

Background: Polycaprolactone (PCL) scaffolds exhibit high biocompatibility and are attractive as vascular conduits. Materials & methods: PCL tubes were cultivated in bioreactor with human adipose regenerative cells to assess ex vivo cytocompatibility, whereas in vivo PCL tube patency was evaluated in sheep carotid bypass with and without antithrombotic treatment. Results:Ex vivo results revealed increasing adipose regenerative cells on PCL using dynamic bioreactor culturing. In vivo data showed that 67% (2/3) of grafts in the antithrombotic group were patent at day 28, while 100% (3/3) of control grafts were occluded already during the first week due to thrombosis. Histology showed that patent PCL grafts were recellularized by host cells. Conclusion: PCL tubes may work as small diameter vascular scaffolds under antithrombotic treatment.

Adipose-derived regenerative cells and lipotransfer in alleviating breast cancer-related lymphedema: An open-label phase I trial with 4 years of follow-up.

Patients with breast cancer-related lymphedema (BCRL) have reduced quality of life and arm function. Current treatments are palliative, and treatments improving lymphedema are lacking. Preclinical studies have suggested that adipose-derived regenerative cells (ADRCs) can alleviate lymphedema. We, therefore, aimed to assess whether ADRCs can alleviate lymphedema in clinical reality with long-term follow-up. We treated 10 patients with BCRL using ADRCs and a scar-releasing lipotransfer to the axillary region, and all patients were followed 1, 3, 6, 12, and 48 months after treatment. The primary endpoint was change in arm volume. Secondary endpoints were safety, change in lymphedema symptoms, quality of life, lymphedema-associated cellulitis, and conservative treatment use. There was no significant decrease in BCRL volume after treatment. However, self-reported upper extremity disability and arm heaviness and tension improved. Six patients reduced their use of conservative BCRL treatment. Five patients felt that their BCRL had improved substantially, and four of these would redo the treatment. We did not observe any cases of locoregional breast cancer recurrence. In this phase I study with 4 years of follow-up, axillary delivered ADRCs and lipotransfer were safe and feasible and improved BCRL symptoms and upper extremity function. Randomized controlled trials are needed to confirm the results of this study.

Injectable and self-healing hydrogel as a stem cells carrier for treatment of diabetic erectile dysfunction.

Diabetic erectile dysfunction has witnessed extensive preclinical and clinical explorations of intracavernous injection of stem cells therapy. However, intracavernous injection of stem cells for diabetic erectile dysfunction is challenged by rapid diffusion from cavernous sinus. Here, we found that a benzaldehyde terminated poly (ethylene glycol)/glycol chitosan (CHO-PEG/GCS) hydrogel with injectability and self-healability served as a stem cell carrier to prolong cell retention in corpus cavernosum. It was able to gelate under physiological condition and encapsulate adipose stem cells (ASCs) without reducing proliferation after injection. Encapsulated labelled ASCs presented higher fluorescence than non-encapsulated ones in the region of penis at 14 days after intracavernous injection in male rats. CHO-PEG/GCS hydrogel enhanced ASCs to ameliorate diabetes-induced fibrosis and apoptosis of CD31-positive endothelial cells, α-SMA-positive smooth muscle and NeuN-positive neural fibers 12 weeks post-operation. It also synergized with ASCs to elevate cGMP level and promote erectile function. CHO-PEG/GCS hydrogel serves as a promising stem cell carrier in conditions requiring injection and in situ gelation to prolong cell retention.

A 12-Month Follow-up After a Single Intracavernous Injection of Autologous Adipose-Derived Regenerative Cells in Patients with Erectile Dysfunction Following Radical Prostatectomy: An Open-Label Phase I Clinical Trial.

OBJECTIVE: To explore safety in adipose-derived regenerative cells (ADRC) therapy, treating erectile dysfunction (ED). METHODS: Twenty-one patients with ED after radical prostatectomy, with no signs of recovery using conventional therapy, received a single intracavernous injection of autologous ADRC and were followed for 1 year. Six men were incontinent, and 15 were continent at inclusion. The primary (safety of ADRC therapy) and secondary endpoints (sexual function) were evaluated at 1, 3, 6, and 12 months after ADRC injection by registration of adverse events and validated questionnaires using the international index of erectile function-5 and erection hardness score. RESULTS: No serious adverse events occurred, but 8 reversible minor events related to the liposuction were noted. Eight out of 15 (53%) patients in the continent group reported erectile function sufficient for intercourse at 12 months. Baseline median international index of erectile function-5 scores (6.0; interquartile range [IQR] 3) were unchanged 1 month after the treatment, but significantly increased after 6 to 7 (IQR 17). This effect was sustained at 12 months (median 8; IQR 14). We did not see any improvements in erectile function in the group of incontinent men or among men with ED prior to radical prostatectomy. CONCLUSION: Intracavernous injection of ADRC is safe in this phase 1 study with a 12-month follow-up.

Human and Autologous Adipose-derived Stromal Cells Increase Flap Survival in Rats Independently of Host Immune Response.

INTRODUCTION: There is a rising interest in adipose-derived stromal cells for clinical use; however, it is unknown whether freshly isolated stromal cells (SVF) or culture-expanded cells (ASCs) are more efficacious. We therefore aimed to compare the 2 cellular therapies in an in vivo model of angiogenesis, the ischemic flap in rats, which induces acute ischemia. We also aimed to determine the importance of cell presence and the host immune response. METHODS: A total of 96 rats (n = 12 in each group) were used, and in each rat, a caudally based random flap measuring 2 × 7 cm was made. The study was conducted in 3 phases. First, each rat was treated with human SVF cells, human ASCs, or vehicle. Second, each rat was treated with human SVF, human SVF lysate, or vehicle. Finally, each rat was treated with rat (autologous) SVF cells or vehicle. Flap survival, vessel density, and stromal cell retention were evaluated after 7 days. RESULTS: The mean survival rates for SVF treatment regardless of human or autologous origin were significantly increased as compared with the control group. Adipose stem/stromal cell and SVF lysate injection did not increase flap survival. Vessel density was increased for human and rat SVF and human ASC but not for SVF lysate. Human cells were not detected in the flaps after 7 days. CONCLUSIONS: Flap survival increased with SVF treatment regardless of human or autologous origin, suggesting that increased flap survival is independent of the host immune response. All cell injections lead to increased vessel density, but it did not necessarily lead to increased flap survival. Further research should elaborate which molecular events make SVF treatment more efficacious than ASC.

Concise Review: A Safety Assessment of Adipose-Derived Cell Therapy in Clinical Trials: A Systematic Review of Reported Adverse Events.

The popularity of adipose-derived cell therapy has increased over the last decade, and the number of studies published annually is growing. However, concerns regarding safety in the setting of previous malignancy or the use of allogeneic cells have been raised. We therefore aimed to systematically review all clinical studies using adipose-derived cell therapy to identify reported adverse events with a special focus on risk of thromboembolic, immunological, and oncological safety concerns. Our systematic search resulted in 70 included studies involving more than 1,400 patients that were treated with adipose-derived cell therapy. Safety assessment method was not described in 32 of the included studies. For studies involving systemic or cardiac administration, one case of pulmonary thromboembolism and cases of both myocardial and cerebral infarctions were described. In the setting of allogeneic cell therapy studies, where the production of specific antibodies toward donor cells was examined, it was noted that 19%-34% of patients develop antibodies, but the consequence of this is unknown. With regard to oncological safety, only one case of breast cancer recurrence was identified out of 121 patients. Adipose-derived cell therapy has so far shown a favorable safety profile, but safety assessment description has, in general, been of poor quality, and only adverse events that are looked for will be found. We encourage future studies to maintain a strong focus on the safety profile of cell therapy, so its safeness can be confirmed. Stem Cells Translational Medicine 2017;6:1786-1794.

Treatment of Breast Cancer-Related Lymphedema with Adipose-Derived Regenerative Cells and Fat Grafts: A Feasibility and Safety Study.

Breast cancer-related lymphedema (BCRL) is a debilitating late complication with a lack of treatment opportunities. Recent studies have suggested that mesenchymal stromal cells can alleviate lymphedema. Herein, we report the results from the first human pilot study with freshly isolated adipose-derived regenerative cells (ADRC) for treating lymphedema with 6 months follow-up. Ten BCRL patients were included. ADRC was injected directly into the axillary region, which was combined with a scar-releasing fat graft procedure. Primary endpoints were change in arm volume. Secondary endpoints were change in patient reported outcome and safety. The study is registered with ClinicalTrials.gov (NCT02592213). During follow-up, a small volume reduction was noted but was not significant. Five patients reduced their use of conservative management. Patient-reported outcomes improved significantly over time. ADRCs were well tolerated and only minor transient adverse events related to liposuction were noted. In this pilot study, a single injection of ADRC improved lymphedema based on patient-reported outcome measures, and there were no serious adverse events in the 6 months follow-up period. In addition, half of the patients reduced their use of conservative management. ADRC therapy is a promising interventional therapy for alleviating lymphedema, but results need to be confirmed in randomized clinical trials. Stem Cells Translational Medicine 2017;6:1666-1672.

The non-canonical NOTCH1 ligand Delta-like 1 homolog (DLK1) self interacts in mammals.

Delta-like 1 homolog (DLK1) is an imprinted gene, which is widely expressed during mammalian development and plays a pivotal role in differentiation of various tissue types. Most recently, we have shown that DLK1 interacts with NOTCH1, yet several Notch independent mechanisms have previously been suggested as well, but only poorly confirmed in a mammalian context. In the present study, we employed the mammalian two-hybrid (MTH) system, a genetic in vivo protein-protein interaction system, to show robust DLK1-DLK1, DLK1-FnI (Fibronectin) and DLK1-CFR (cysteine-rich FGF receptor) interactions, whereas the proposed DLK1-IGFBP1 interaction was not supported by MTH. Very little has previously been described on the DLK1 self-interaction. Herein, we showed by immunoprecipitation as well as Sulfo-SBED label transfer that the DLK1-DLK1 interaction likely is part of Dlk1's function in preadipocytes. Furthermore our data suggest that DLK1 interacts with itself through EGF domain 4 and 5, which is distinct from the recently described NOTCH1-DLK1 interaction, which occurs between EGF domain 5 and 6. This opens up the possibility that Notch independent mechanisms like the DLK1-DLK1 interaction may modulate the non-canonical NOTCH1-DLK1 interaction further complexing this system.

A 3-month age difference profoundly alters the primary rat stromal vascular fraction phenotype.

The stromal vascular fraction (SVF) is a heterogeneous population obtained from collagenase digestion of adipose tissue. When cultured the population becomes more homogeneous and the cells are then termed adipose stromal/stem cells (ASCs). Both the freshly isolated primary SVF population and the cultured ASC population possess regenerative abilities suggested to be mediated by paracrine mechanisms mainly. The use of ASCs and SVF cells, both in animal studies and human clinical studies, has dramatically increased during recent years. However, more knowledge regarding optimal donor characteristics such as age is demanded. Here we report that even a short age difference has an impact on the phenotype of primary SVF cells. We observed that a 3-month difference in relatively young adult rats affects the expression pattern of several mesenchymal stem cell markers in their primary SVF. The younger animals had significantly more CD90+/CD44+/CD29+/PDGFRα+primary cells, than the aged rats, suggesting an age dependent shift in the relative cell type distribution within the population. Taken together with recent studies of much more pronounced age differences, our data strongly suggest that donor age is a very critical parameter that should be taken into account in future stem cell studies, especially when using primary cells.

Cell-Assisted Lipotransfer Using Autologous Adipose-Derived Stromal Cells for Alleviation of Breast Cancer-Related Lymphedema.

UNLABELLED: : Lymphedema is one of the most frequent side effects following cancer treatment, and treatment opportunities for it are currently lacking. Stem cell therapy has been proposed as a possible novel treatment modality. This study was the first case in which freshly isolated adipose-derived stromal cells were used to treat lymphedema. Treatment was given as a cell-assisted lipotransfer in which 4.07 × 10(7) cells were injected with 10 ml of lipoaspirate in the axillary region. Four months after treatment, the patient reported a great improvement in daily symptoms, reduction in need for compression therapy, and volume reduction of her affected arm. There were no adverse events. The outcome for this patient provides support for the potential use of cellular therapy for lymphedema treatment. We have begun a larger study to further test the feasibility and safety of this procedure (ClinicalTrials.gov Identifier NCT02592213). SIGNIFICANCE: Lymphedema is a very debilitating side effect of cancer treatment and has very few treatment options. Stem cell therapy has the potential to change the treatment paradigm from a conservative to a more curative approach. Freshly isolated, autologous, adipose-derived stromal cells were combined with a fat-graft procedure to treat lymphedema. The treated patient had great improvement in daily symptoms, a reduced need for compression therapy, and a reduction in arm volume after 4 months. There were no adverse events. The use of cellular therapy for lymphedema treatment is supported by this patient's outcome. A phase II study has begun to further test its feasibility and safety.

Safety and Potential Effect of a Single Intracavernous Injection of Autologous Adipose-Derived Regenerative Cells in Patients with Erectile Dysfunction Following Radical Prostatectomy: An Open-Label Phase I Clinical Trial.

BACKGROUND: Prostate cancer is the most common cancer in men, and radical prostatectomy (RP) often results in erectile dysfunction (ED) and a substantially reduced quality of life. The efficacy of current interventions, principal treatment with PDE-5 inhibitors, is not satisfactory and this condition presents an unmet medical need. Preclinical studies using adipose-derived stem cells to treat ED have shown promising results. Herein, we report the results of a human phase 1 trial with autologous adipose-derived regenerative cells (ADRCs) freshly isolated after a liposuction. METHODS: Seventeen men suffering from post RP ED, with no recovery using conventional therapy, were enrolled in a prospective phase 1 open-label and single-arm study. All subjects had RP performed 5-18 months before enrolment, and were followed for 6 months after intracavernosal transplantation. ADRCs were analyzed for the presence of stem cell surface markers, viability and ability to differentiate. Primary endpoint was the safety and tolerance of the cell therapy while the secondary outcome was improvement of erectile function. Any adverse events were reported and erectile function was assessed by IIEF-5 scores. The study is registered with ClinicalTrials.gov, NCT02240823. FINDINGS: Intracavernous injection of ADRCs was well-tolerated and only minor events related to the liposuction and cell injections were reported at the one-month evaluation, but none at later time points. Overall during the study period, 8 of 17 men recovered their erectile function and were able to accomplish sexual intercourse. Post-hoc stratification according to urinary continence status was performed. Accordingly, for continent men (median IIEFinclusion = 7 (95% CI 5-12), 8 out of 11 men recovered erectile function (IIEF6months = 17 (6-23)), corresponding to a mean difference of 0.57 (0.38-0.85; p = 0.0069), versus inclusion. In contrast, incontinent men did not regain erectile function (median IIEF1/3/6 months = 5 (95% CI 5-6); mean difference 1 (95% CI 0.85-1.18), p > 0.9999). INTERPRETATION: In this phase I trial a single intracavernosal injection of freshly isolated autologous ADRCs was a safe procedure. A potential efficacy is suggested by a significant improvement in IIEF-5 scores and erectile function. We suggest that ADRCs represent a promising interventional therapy of ED following prostatectomy. FUNDING: Danish Medical Research Council, Odense University Hospital and the Danish Cancer Society.

Persistent scarring and dilated cardiomyopathy suggest incomplete regeneration of the apex resected neonatal mouse myocardium--A 180 days follow up study.

Heart damage in mammals is generally considered to result in scar formation, whereas zebrafish completely regenerate their hearts following an intermediate and reversible state of fibrosis after apex resection (AR). Recently, using the AR procedure, one-day-old mice were suggested to have full capacity for cardiac regeneration as well. In contrast, using the same mouse model others have shown that the regeneration process is incomplete and that scarring still remains 21 days after AR. The present study tested the hypothesis that like in zebrafish, fibrosis in neonatal mammals could be an intermediate response before the onset of complete heart regeneration. Myocardial damage was performed by AR in postnatal day 1 C57BL/6 mice, and myocardial function and scarring assessed at day 180 using F-18-fluorodeoxyglucose positron emission tomography (FDG-PET) and histology, respectively. AR mice exhibited decreased ejection fraction and wall motion with increased end-diastolic and systolic volumes compared to sham-operated mice. Scarring with collagen accumulation was still substantial, with increased heart size, while cardiomyocyte size was unaffected. In conclusion, these data thus show that apex resection in mice results in irreversible fibrosis and dilated cardiomyopathy suggesting that cardiac regeneration is limited in neonatal mammals and thus distinct from the regenerative capacity seen in zebrafish.