IBRORS-MCL study: a Spanish retrospective and observational study of relapsed/refractory mantle-cell lymphoma treated with ibrutinib in routine clinical practice.

This retrospective study evaluated 66 patients diagnosed with relapsed and/or refractory mantle cell lymphoma (R/R MCL) treated with ibrutinib in Spain in routine clinical practice. At diagnosis, patients had a median age of 64.5 years, 63.6% presented with intermediate/high sMIPI (simplified prognostic index for advanced-stage mantle cell lymphoma), 24.5% had the blastoid variant, and 55.6% had a Ki67 > 30%. Patients had received a median of 2 prior lines of therapy (range 1-2; min-max 1-7). Overall response rate was 63.5%, with 38.1% of patients achieving complete response (CR). With a median duration of ibrutinib exposure of 10.7 months (range 5.2-19.6; min-max 0.3-36), the median progression-free survival (PFS) and overall survival (OS) were 20 months [95% confidence interval (CI) 8.8-31.1] and 32 months (95% CI 22.6-41.3), respectively, and were not reached in patients achieving CR. No grade ≥ 3 cardiovascular toxicity or bleeding was reported. This study supports that treatment with ibrutinib leads to high response rates and favorable survival outcomes in patients with R/R MCL.

Clinical grade production of IL-15 stimulated NK cells for early infusion in adult AML patients undergoing haploidentical stem cell transplantation with post-transplant cyclophosphamide.

BACKGROUND: Allogeneic stem cell transplantation is the treatment of choice for acute myeloid leukemia (AML) patients. Unmanipulated haploidentical transplantation (Haplo-HSCT) is commonly used for those AML patients who need a timely transplant and do not have a suitable matched donor, but relapse rates are still high, and improvements are needed. Adoptive immunotherapy using natural killer cells (NK cells) could be a promising tool to improved Haplo-HSCT but, to date, no optimal infusion and manufacturing protocols have been developed. STUDY DESIGN AND METHODS: In this study, we describe a quick and reproducible protocol for clinical-grade production of haploidentical donor NK cells using double immunomagnetic depletion and enrichment protocol and overnight IL-15 stimulation. RESULTS: Thus, we have obtained 8 viable and functional NK cell products that have been safely infused to five AML patients undergoing unmanipulated Haplo-HSCT. DISCUSSION: Our results demonstrate the safety and feasibility of manufactured NK IL15 cells obtained from an adult allogeneic donor in the setting of haploidentical transplantation for AML patients.

Adult Cells Combined With Platelet-Rich Plasma for Tendon Healing: Cell Source Options.

BACKGROUND: The combination of cells with platelet-rich plasma (PRP) may fulfill tendon deficits and help overcome the limited ability of tendons to heal. PURPOSE: To examine the suitability of 3 human cell types in combination with PRP and the potential impact of the tenocyte-conditioned media (CM) to enhance tendon healing. STUDY DESIGN: Controlled laboratory study. METHODS: Tenocytes, bone marrow-derived mesenchymal stem cells, and skin fibroblasts were cultured in 3-dimensional PRP hydrogels supplemented or not with CM, and cell proliferation and migration were examined. The effect of tendon-derived CM on matrix-forming phenotype and secretion of inflammatory proteins was determined through their administration to mesenchymal stem cells, tendon, and skin fibroblasts by reverse transcription quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. RESULTS: Differences were found in the matrix-forming phenotype between each of the cell types. The ratio of collagen I:collagen III was greater in bone marrow-derived mesenchymal stem cells than in skin fibroblasts and tenocytes. The bone marrow-derived mesenchymal stem cells expressed increased levels of cartilage-related genes than tenocytes or skin fibroblasts. The presence of the tenocyte-CM stimulated basic healing mechanisms including proliferation and chemotaxis in all cell types. In addition, the tenocyte-CM modified the matrix-forming phenotype of every cell type when cultured in PRP hydrogels. Each cell type secreted interleukin-6, interleukin-8, and monocyte chemotactic protein-1 in PRP hydrogels, but mesenchymal stem cells secreted less interleukin-8 and monocyte chemotactic protein-1 than tenocytes or skin fibroblasts. CONCLUSION: The tenocyte-CM combined with PRP stimulated tenogenesis in mesenchymal stem cells and in skin fibroblasts and reduced the secretion of inflammatory proteins. CLINICAL RELEVANCE: Modifying the target tissue with PRP prior to cell implantation may optimize the effect of cell therapies. Skin fibroblasts and bone marrow-derived mesenchymal stem cells combined with PRP could be used to regenerate tendons.

Platelet Rich Plasma and Culture Configuration Affect the Matrix Forming Phenotype of Bone Marrow Stromal Cells.

We aim to examine the influence of platelet rich plasma (PRP) and spatial cues in cartilage/bone matrix forming proteins, and to evaluate the mitotic and chemotactic potential of PRP on human mesenchymal stem cells (hMSCs). Directed cell migration towards PRP gradients was assessed in chemotactic chambers, and recorded by time-lapse microscopy. hMSCs cultured in three-dimensional (3D) scaffolds were visualized by scanning electron microscopy; Hoechst dye was used to confirm cell confluence in 3D-constructs and monolayers before experimental treatment. MSCs were treated with 10% PRP lysate or 10% PRP lysate supplemented with TGF-ß-based differentiation medium. The expression of cartilage (COL2A1, Sox9, ACAN, COMP), and bone (COL1A1, VEGF, COL10A1, Runx2) fundamental genes was assessed by real time PCR in monolayers and 3D-constructs. PRP had mitotic (p < 0.001), and chemotactic effect on hMSCs, Ralyleigh test p = 1.02E - 10. Two and three-week exposure of MSCs to PRP secretome in 3D-constructs or monolayers decreased Sox9 expression (p < 0.001 and p = 0.050) and COL2A1, (p = 0.011 and p = 0.019). MSCs in monolayers exposed to PRP showed increased ACAN (p = 0.050) and COMP (p < 0.001). Adding TGF-ß-based differentiation medium to PRP increased COMP, and COL2A1 expression at gene and protein level, but merely in 3D-constructs, p < 0.001. TGF-ß addition to monolayers reduced Sox9 (p < 0.001), aggrecan (p = 0.004), and VEGF (p = 0.004). Cells exposed to PRP showed no changes in hypertrophy associated genes in either monolayers or 3D-constructs. Our study suggests hMSCs have high-degree of plasticity having the potential to change their matrix-forming phenotype when exposed to PRP and according to spatial configuration.

The Properties of 3 Different Plasma Formulations and Their Effects on Tendinopathic Cells.

BACKGROUND: Tendinopathies are attributed to failure of the healing process and inadequate tissue remodeling. Plasma injections can trigger regenerative responses by modifying the molecular microenvironment. PURPOSE: To examine the differences in the mitotic, chemotactic, anabolic, and inflammatory effects between leukocyte- and platelet-rich plasma (L-PRP), platelet-rich plasma (PRP), and platelet-poor plasma (PPP). STUDY DESIGN: Controlled laboratory study. METHODS: Tendinopathic cells were cultured in 3-dimensional (3D) hydrogels formed using PPP, PRP, and L-PRP. Cell migration was evaluated using a µ-Slide chemotaxis chamber with video microscopy. Proliferation was assessed using XTT assays. Expression of genes associated with matrix turnover, including type 1 collagen (COL1A1), COL3A1, aggrecan, decorin, fibronectin, matrix metalloproteinase 1 (MMP-1), MMP-3, A Disintegrin-Like And Metalloprotease With Thrombospondin Type 1 Motif proteins 4 (ADAMTS-4), and ADAMTS-5, was assessed using real-time reverse-transcription polymerase chain reaction. Secreted inflammatory proteins, including interleukin (IL)-1ß, IL-6, IL-8, monocyte chemotactic protein 1 (MCP-1), and regulated on activation, normal T cell expressed and secreted (RANTES), as well as vascular endothelial growth factor (VEGF) and connective tissue growth factor (CTGF), were quantified using enzyme-linked immunosorbent assay. RESULTS: Tendinopathic cells migrate at a higher velocity along L-PRP and PRP than along PPP gradients. PRP and L-PRP promote hypercellularity. PPP and PRP showed more pronounced anabolic properties, as demonstrated by enhanced COL1A1 and COL3A1 and reduced MMP-1 expression. Decorin, fibronectin, and aggrecan were downregulated in L-PRP compared with PPP and PRP. L-PRP and PRP were shown to be more proinflammatory than PPP in terms of IL-6 secretion, but cells in PPP showed MCP-1(high) phenotype. CTGF secretion was significantly reduced in L-PRP compared with PPP and PRP. CONCLUSION: The main advantages of L-PRP and PRP use, compared with PPP, include their stronger chemotactic and proliferative properties. While PPP and PRP stimulate matrix anabolism, L-PRP is more proinflammatory. Emphasis should be placed on the temporal needs and biological characteristics of injured tendons, and plasma formulations need to be tailored accordingly. CLINICAL RELEVANCE: Versatile systems allowing the preparation of different plasma formulations, such as PPP, PRP, or L-PRP, can help refine clinical applications by taking advantage of their different biological properties.

Biological approach for the management of non-healing diabetic foot ulcers.

OBJECTIVE: To show an approach to profit of the main components of platelet rich plasma (PRP), i.e. the signaling proteins, and the fibrin scaffold and discuss the intervention within TIME (Tissue, Inflammation/Infection, Moisture, Edges) framework. METHODS: Two patients with diabetic foot ulcers are treated with both liquid and gelled PRP, and the rationale for the PRP intervention is described herein. Autologous blood is withdrawn and, PRP is separated by single spinning and activated with CaCl2 prior to application. PRP is injected in an activated liquid form, i.e. freshly activated, before coagulation, within the wound edges. In fibrotic tissue PRP is introduced performing a needling procedure. In addition, PRP, clotted ex-vivo, is applied in the wound bed as a primary dressing. RESULTS: Both patients responded positively to PRP intervention. Case 1 healed after five weekly PRP applications. Case 2 healed after eight weekly PRP applications. Patient satisfaction was high in both cases. The procedure had no complications, is well tolerated and easy to perform in any medical setting. CONCLUSION: PRP intervention is safe and if associated with correct tissue debridement and preparation of the host tissue it may help to decrease the burden of diabetic foot ulcers. Carefully designed randomized clinical trials with special attention to the PRP procedure are needed to assess the efficacy of these interventions.

Three-Dimensional Platelet-Rich Plasma Hydrogel Model to Study Early Tendon Healing.

Since the experimental conditions of cell cultures may bias results, it is critical to use suitable models. This is also true in the context of tendon cell biology and the study of platelet-rich plasma (PRP) therapies and PRP-augmented cell-based therapies. We compared the culture of human tendon cells in 2 dimensions (2D) with PRP-supplemented media to culture in matching 3-dimensional (3D) PRP hydrogels. Cell proliferation, cell shape, and the pattern of gene and protein expression were examined. Our data revealed modifications in cell shape and enhanced expression of tenomodulin and scleraxis in 3D hydrogels. Additionally, protein secretion analysis using glass-based arrays specific for angiogenesis revealed differences in interleukin (IL)-6 and IL-8 protein expression between 2D cultures and 3D hydrogels, while the secretion of other angiogenic or inflammatory cytokines was unaffected. Our study suggests that 3D hydrogels are physiologically more relevant than 2D cultures in the study of tendon cells, based on cell shape, support of tenocyte proliferation, phenotype, and the pattern of gene and protein expression.

Effect of bioactive glass particles on osteogenic differentiation of adipose-derived mesenchymal stem cells seeded on lactide and caprolactone based scaffolds.

Incorporation of bioactive glass (BG) particles to synthetic polymer scaffolds is a promising strategy to improve the bioactivity of bioinert materials and to stimulate specific cell responses. In this study, the influence of incorporating BG particles to lactide and caprolactone based porous scaffolds on osteogenic differentiation of adipose-derived stem cells (ASCs) was analyzed. Accordingly, ASCs were seeded on poly(L-lactide) (PLLA), poly(ε-caprolactone) (PCL), or poly(L-lactide-co-ε-caprolactone) (PLCL) scaffolds containing 15 vol % of BG particles in two culture conditions: standard versus osteogenic culture medium. In standard culture medium, incorporation of BG to a PLLA scaffold increased the ALP activity with respect to its unfilled counterpart (ca. 1.2- and a 1.6-fold increase over 7 and 14 days, respectively). Moreover, in all the studied polymers the incorporation of BG induced a slightly higher production of mineralized matrix by ASCs, but the differences observed were not statistically significant. In the osteogenic medium, the effect of BG was masked by the effect of osteogenic supplements in the long-term. However, in the short-term (day 7), BG particles induced an early ALP activity of predifferentiated osteoblasts on PLLA and PCL scaffolds and higher matrix mineralization on PCL scaffolds. In summary, the addition of BG particles to PLLA and PCL scaffolds sustains ASC osteogenic differentiation, facilitates mineralization and induces the formation of a hydroxyapatite layer on the surface of the polymer scaffolds.

Platelet-rich plasma modulates the secretion of inflammatory/angiogenic proteins by inflamed tenocytes.

BACKGROUND: Platelet-rich plasma therapies for tendinopathy appear to provide moderate pain reduction. However, the biological mechanisms behind the observed clinical effects remain poorly characterized. QUESTIONS/PURPOSES: The purpose of this study was to explore whether platelet-rich plasma modifies the inflammatory/angiogenic status of already inflamed tenocytes by examining (1) gene expression; (2) modulation of chemokine and interleukin secretion; and (3) differences between healthy and tendinopathic tenocytes. METHODS: Cells from both healthy and tendinopathic tendons were exposed to interleukin (IL)-1ß and after treated with platelet-rich plasma. Modifications in the expression of selected genes were assessed by real-time reverse transcription-polymerase chain reaction and changes in secretion of angiogenic/inflammatory molecules by enzyme-linked immunosorbent assay. Platelet-rich plasma-induced changes in tendinopathic cells were compared with normal after normalizing platelet-rich plasma data against IL-1ß status in each specific sample. RESULTS: In IL-1ß-exposed cells, platelet-rich plasma downregulates expression of IL-6/CXCL-6 (mean, 0.015; 95% confidence interval [CI], 0.005-0.025; p = 0.026), IL-6R (mean, 0.61; 95% CI, 0.27-0.95; p = 0.029), and IL-8/CXCL-8 (mean, 0.02; 95% CI, 0.007-0.023; p = 0.026). Secretion of IL-6/CXCL6, 0.35 (95% CI, 0.3-0.4; p = 0.002), IL-8/CXCL8, 0.55 (95% CI, 0.5-0.7; p = 0.01), and monocyte chemoattractant protein-1/CCL2, 0.40 (95% CI, 0.2-0.6; p = 0.001) was reduced by platelet-rich plasma, whereas vascular endothelial growth factor increased by twofold, (95% CI, 1.7-2.3; p < 0.001). RANTES/CCL5 increased by10-fold (95% CI, 4-17) and hepatocyte growth factor by 21-fold (95% CI, 0.2-42) in tendinopathic and by 2.3-fold (95% CI, 2-3) and threefold (95% CI, 1-5) in normal cells (p = 0.005 for both). CONCLUSIONS: Platelet-rich plasma induces an immunomodulatory and proangiogenic phenotype consistent with healing mechanisms with few differences between tendinopathic and normal cells. CLINICAL RELEVANCE: Platelet-rich plasma injections in pathological and nearby tissue might help to recover tendon homeostasis.

Angiogenic and innate immune responses triggered by PRP in tendon cells are not modified by hyperuricemia.

BACKGROUND: hyperuricemia is becoming a critical medical problem, and a current focus of research. Uric acid is also a death cell associated stressor that may trigger innate immune responses via the synthesis of inflammatory and angiogenic proteins. PURPOSE: to investigate the angiogenic/inflammatory protein profile of tendon cells treated with Platelet Rich Plasma (PRP), and to assess if there are any differences in synthesis of angiogenic/inflammatory cytokines between PRP-treated or hyperuricemic PRP-treated cells. METHODS: tendon cells were treated with PRP or hyperuricemic PRP and cell culture supernatants examined using glass based protein arrays for inflammation and angiogenesis. Relevant proteins were subsequently quantified by ELISA or EASIA methods. RESULTS: the impact of PRP on angiogenesis and inflammation is evidenced by relevant cytokine synthesis including: Monocyte Chemoattractant Protein (MCP-1/CCL2), Regulated upon Activation Normally T cells Expressed and Secreted (RANTES/CCL5), IL-6/CXCL6, IL-8/CXCL8, Vascular Endothelial Growth Factor (VEGF), Growth Regulated Protein (GRO-a/CXCL1) and Hepatocyte Growth Factor (HGF). IL-1beta was not detected in these conditions. Taken together these data suggest an initial angiogenetic and innate immune responses driven by chemokines that is not altered by the presence of hyperuricemia, at this point, except for IL-8 secretion, p= 0.042.

Partnership between platelet-rich plasma and mesenchymal stem cells: in vitro experience.

We aim to identify current in vitro research exploring platelet-rich plasma (PRP) effects in human Mesenchymal Stem Cells (MSCs) that may encourage or limit the clinical application of MSCs along with PRP. After a systematic search, we identified 57 in vitro studies, focused on optimization of MSC manufacturing, and expanding knowledge about how PRP modifies MSCs behavior for translational purposes. Influences of PRP on proliferation, migration, stemness, preservation of MSC immune-modulatory properties and appearance of senescence phenotype have been explored. Overall PRP stimulates MSC proliferation, preserves MSCs multipotency and does not interfere with any lineage differentiation. PRP (as platelet lysate or releasate) preserves the immune-privileged potential of MSCs and may delay the appearance of the senescent phenotype. Currently there are few data linking precise molecules and biological mechanisms. Various gaps of knowledge need to be addressed in order to obtain enough useful information for translational purposes.

Biocompatible bacterial cellulose-poly(2-hydroxyethyl methacrylate) nanocomposite films.

A series of bacterial cellulose-poly(2-hydroxyethyl methacrylate) nanocomposite films was prepared by in situ radical polymerization of 2-hydroxyethyl methacrylate (HEMA), using variable amounts of poly(ethylene glycol) diacrylate (PEGDA) as cross-linker. Thin films were obtained, and their physical, chemical, thermal, and mechanical properties were evaluated. The films showed improved translucency compared to BC and enhanced thermal stability and mechanical performance when compared to poly(2-hydroxyethyl methacrylate) (PHEMA). Finally, BC/PHEMA nanocomposites proved to be nontoxic to human adipose-derived mesenchymal stem cells (ADSCs) and thus are pointed as potential dry dressings for biomedical applications.