The species-specific regenerative effects of notochordal cell-conditioned medium on chondrocyte-like cells derived from degenerated human intervertebral discs

During intervertebral disc (IVD) maturation, the main cell type shifts from notochordal cells (NCs) to chondrocytelike cells (CLCs). NCs secrete factors with regenerative potential, making them an interesting focus for regenerative treatments. During initial development, these strategies preferably employ non-human donors due to easy availability of their NC-rich nucleus pulposus (NP) tissue. To increase the success of translating these strategies for clinical application, this study aimed to delineate whether NC-secreted factors of different species have a regenerative effect on human CLCs. Human, canine and porcine NC-rich NP tissue and NC-conditioned medium (NCCM) were analysed biochemically and histologically. Human CLC micro-aggregates from degenerated IVDs were cultured in human, canine or porcine NCCM. Collagen, glycosaminoglycan (GAG) and DNA content was determined and histology was performed. Canine and porcine NPs were richer in NCs than human NPs. Human NPs contained the highest collagen content, whereas the DNA and GAG content of canine NPs was significantly higher than that of human or porcine NPs. NCCM from all species significantly increased the DNA and GAG content of the human CLC micro-aggregates. Porcine and canine NCCM were significantly more potent than human NCCM in inducing GAG deposition, whereas only human NCCM induced collagen type II production. Secreted factors from human, canine and porcine NC-rich NPs exerted regenerative effects on human CLCs, indicating a cross-species effect. Bioactive compound(s) are present in NCCM of different species that may reverse human IVD degeneration, supporting further research into strategies based on NC-technology employing canine or porcine models for their translation into humans.</p

What does cell therapy manufacturing cost? A framework and methodology to facilitate academic and other small-scale cell therapy manufacturing costings

Background aims: Recent technical and clinical advances with cell-based therapies (CBTs) hold great promise in the treatment of patients with rare diseases and those with high unmet medical need. Currently the majority of CBTs are developed and manufactured in specialized academic facilities. Due to small scale, unique characteristics and specific supply chain, CBT manufacturing is considered costly compared to more conventional medicinal products. As a result, biomedical researchers and clinicians are increasingly faced with cost considerations in CBT development. The objective of this research was to develop a costing framework and methodology for academic and other small-scale facilities that manufacture cell-based therapies. Methods: We conducted an international multi-center costing study in four facilities in Europe using eight CBTs as case studies. This study includes costs from cell or tissue procurement to release of final product for clinical use. First, via interviews with research scientists, clinicians, biomedical scientists, pharmacists and technicians, we designed a high-level costing framework. Next, we developed a more detailed uniform methodology to allocate cost items. Costs were divided into steps (tissue procurement, manufacturing and fill-finish). The steps were each subdivided into cost categories (materials, equipment, personnel and facility), and each category was broken down into facility running (fixed) costs and operational (variable) costs. The methodology was tested via the case studies and validated in developer interviews. Costs are expressed in 2018 euros (€). Results: The framework and methodology were applicable across facilities and proved sensitive to differences in product and facility characteristics. Case study cost estimates ranged between €23 033 and €190 799 Euros per batch, with batch yield varying between 1 and 88 doses. The cost estimations revealed hidden costs to developers and provided insights into cost drivers to help design manufacturing best practices. Conclusions: This framework and methodology provide step-by-step guidance to estimate manufacturing costs specifically for cell-based therapies manufactured in academic and other small-scale enterprises. The framework and methodology can be used to inform and plan cost-conscious strategies for CBTs

Development of a value-based healthcare burns core set for adult burn care

Background: Value-based healthcare (VBHC) is increasingly implemented in healthcare worldwide. Transparent measurement of the outcomes most important and relevant to patients is essential in VBHC, which is supported by a core set of most important quality indicators and outcomes. Therefore, the aim of this study was to develop a VBHC-burns core set for adult burn patients. Methods: A three-round modified national Delphi study, including 44 outcomes and 24 quality indicators, was conducted to reach consensus among Dutch patients, burn care professionals and researchers. Items were rated on a nine-point Likert scale and selected if ≥ 70% in each group considered an item ‘important’. Subsequently, instruments quantifying selected outcomes were identified based on a literature review and were chosen in a consensus meeting using recommendations from the Dutch consensus-based standard set and the Dutch Centre of Expertise on Health Disparities. Time assessment points were chosen to reflect the burn care and patient recovery process. Finally, the initial core set was evaluated in practice, leading to the adapted VBHC-burns core set. Results: Twenty-seven patients, 63 burn care professionals and 23 researchers participated. Ten outcomes and four quality indicators were selected in the Delphi study, including the outcomes pain, wound healing, physical activity, self-care, independence, return to work, depression, itching, scar flexibility and return to school. Quality indicators included shared decision-making (SDM), the number of patients receiving aftercare, determination of burn depth, and assessment of active range of motion. After evaluation of its use in clinical practice, the core set included all items except SDM, which are assessed by 9 patient-reported outcome instruments or measured in clinical care. Assessment time points included are at discharge, 2 weeks, 3 months, 12 months after discharge and annually afterwards. Conclusion: A VBHC-burns core set was developed, consisting of outcomes and quality indicators that are important to burn patients and burn care professionals. The VBHC-burns core set is now systemically monitored and analysed in Dutch burn care to improve care and patient relevant outcomes. As improving burn care and patient relevant outcomes is important worldwide, the developed VBHC-burns core set could be inspiring for other countries.</p

Child abuse and neglect in paediatric burns: The majority is caused by neglect and thus preventable

Introduction: It is complex to distinguish negligent burns from inflicted and non-intentional burns, especially since no deliberate action caused the burn. Its recognition is important to create a safe life without (future) burns for our loved-ones who cannot take care of themselves (yet). Our aim was to investigate the incidence and associated characteristics of negligent burns are among children treated at a burn center. Methods: We retrospectively reviewed medical files of all children attending a Dutch burn center with an acute burn between January 2013–December 2015. The conclusion of the Child Abuse and Neglect team (CAN) was used to define inflicted, negligent or non-intentional burns. The remaining cases were reviewed by an expert panel using a standardized protocol. Demographic and social data as well as clinical information about the burn were collected. Results: In total 330 children were included. Negligent burns accounted for 56%, non-intentional for 42% and inflicted for 2% of the burns. Negligent burns were associated with: hot beverages (OR 4.40, 95%CI 2.75–7.05), a younger age (p < 0.001), occurrence at home (OR 4.87 95%CI 2.80–8.45) and were located at the anterior trunk (OR 2.75, 95%CI 1.73–4.35) and neck (OR 1.98, 95%CI 1.12–3.50). Conclusion: This study shows that neglect is a major factor in the occurrence of burns in young children, therefore we conclude that the majority of paediatric burns are preventable. Educational programs creating awareness and focussing on prevention of these burns should be aimed at households, since the majority of negligent as cause of preventable burns occur at home

Bacterial Species and Load Increase During Negative Pressure Wound Therapy: A Prospective Cohort Study

Introduction. The course of both the bacterial species and load and the incidence of infection during negative pressure wound therapy (NPWT) are unclear, with published studies presenting contradicting results. Objective. The aim of the study is to assess the changes in both bacterial species and load, as well as the incidence of infection, before and after NPWT in a patient population with a variety of wounds. Methods. Surgical patients 18 years of age or older who needed NPWT were included in this multicenter, prospective cohort study. A wound swab culture was taken before NPWT and either immediately following NPWT or 6 weeks of follow-up. The change of bacterial species, bacterial load, and rate of infection were determined before and after the start of NPWT. Results. In total, 104 patients were analyzed. The number of positive cultures increased from pre- to post-NPWT. The most cultured pathogenic bacterium was Staphylococcus aureus. The bacterial load was moderately higher at the end of NPWT than at the start (P < .0001). It was noted that 2 swabs contained multidrug-resistant bacteria, 1 pre-NPWT and 1 post-NPWT. Prior to NPWT, 26 patients had a wound infection, 5 of which had a persisting infection at the end of the study. Post-NPWT, 14 patients developed a wound infection. Conclusions. The number of S aureus strains and overall bacterial load increased during NPWT, and the incidence of infection remained the same. Further studies should be conducted to determine whether the increase in bacterial load influences other wound outcome parameters.Trauma Surger

The Stimulatory Effect of Notochordal Cell-Conditioned Medium in a Nucleus Pulposus Explant Culture

Objectives: Notochordal cell-conditioned medium (NCCM) has previously shown to have a stimulatory effect on nucleus pulposus cells (NPCs) and bone marrow stromal cells (BMSCs) in alginate and pellet cultures. These culture methods provide a different environment than the nucleus pulposus (NP) tissue, in which the NCCM ultimately should exert its effect. The objective of this study is to test whether NCCM stimulates NPCs within their native environment, and whether combined stimulation with NCCM and addition of BMSCs has a synergistic effect on extracellular matrix production. Methods: Bovine NP tissue was cultured in an artificial annulus in base medium (BM), porcine NCCM, or BM supplemented with 1 μg/mL Link N. Furthermore, BM and NCCM samples were injected with 106 BMSCs per NP sample. Samples were cultured for 4 weeks, and analyzed for biochemical contents (water, glycosaminoglycan [GAG], hydroxyproline, and DNA), gene expression (COL1A1, COL2A1, ACAN, and SOX9), and histology by Safranin O/Fast Green staining. Results: Culture in NCCM resulted in increased proteoglycan content compared to day 0 and BM, similar to Link N. However, only minor differences in gene expression compared to day 0 were observed. Addition of BMSCs did not result in increased GAG content, and surprisingly, DNA content in BMSC-injected groups was not higher than in the other groups after 4 weeks of culture. Discussion: This study shows that, indeed, NCCM is capable of stimulating NPC matrix production within the NP environment. The lack of increased DNA content in the BMSC-injected groups indicates that BMSCs have died over time. Identification of the bioactive factors in NCCM is crucial for further development of an NCCM-based treatment for intervertebral disc regeneration

The species-specific regenerative effects of notochordal cell-conditioned medium on chondrocyte-like cells derived from degenerated human intervertebral discs

During intervertebral disc (IVD) maturation, the main cell type shifts from notochordal cells (NCs) to chondrocyte-like cells (CLCs). NCs secrete factors with regenerative potential, making them an interesting focus for regenerative treatments. During initial development, these strategies preferably employ non-human donors due to easy availability of their NC-rich nucleus pulposus (NP) tissue. To increase the success of translating these strategies for clinical application, this study aimed to delineate whether NC-secreted factors of different species have a regenerative effect on human CLCs. Human, canine and porcine NC-rich NP tissue and NC-conditioned medium (NCCM) were analysed biochemically and histologically. Human CLC micro-aggregates from degenerated IVDs were cultured in human, canine or porcine NCCM. Collagen, glycosaminoglycan (GAG) and DNA content was determined and histology was performed. Canine and porcine NPs were richer in NCs than human NPs. Human NPs contained the highest collagen content, whereas the DNA and GAG content of canine NPs was significantly higher than that of human or porcine NPs. NCCM from all species significantly increased the DNA and GAG content of the human CLC micro-aggregates. Porcine and canine NCCM were significantly more potent than human NCCM in inducing GAG deposition, whereas only human NCCM induced collagen type II production. Secreted factors from human, canine and porcine NC-rich NPs exerted regenerative effects on human CLCs, indicating a cross-species effect. Bioactive compound(s) are present in NCCM of different species that may reverse human IVD degeneration, supporting further research into strategies based on NC-technology employing canine or porcine models for their translation into humans

Notochordal cell matrix: An inhibitor of neurite and blood vessel growth?

Blood vessel and neurite ingrowth into the degenerating intervertebral disc (IVD) are related to pain. In reported studies, notochordal cell (NC)-conditioned medium (NCCM) induced a regenerative response of nucleus pulposus (NP) cells, but also inhibition of neurite and vessel formation. NC matrix (NCM) derived from NC-rich NP tissue, induced even stronger anabolic effects than NCCM. Thus, the aim was to investigate whether NCM has similar anti-neurogenic and -angiogenic properties as NCCM. NCM and NCCM where produced from porcine NC-rich NP tissue. Human umbilical vein endothelial cells (HUVECs) were cultured in base medium (BM, 300 mOsm), NCCM (produced at 300 and 400 mOsm), NCM, or with chondroitin sulfate (CS, positive control) in angiogenesis-inducing medium, after which vessel length was measured. Although CS alone inhibited vessel growth, NCCM (both osmolarities) stimulated vessel formation by HUVECs. NCM did not affect vessel growth relative to BM. SH-SY5Y cells were cultured in BM, NCCM, and NCM on poly-D-lysine coated and polystyrene surfaces, and analyzed for neurite length and percentage of neurite expressing cells. On coated surfaces, neither NCCM nor NCM affected neurite growth. On a polystyrene surface, NCCM and NCM induced a higher number of neurite-expressing cells. NCCM's previously reported anti-angiogenic and -neurogenic effects were not observed in this study. Although addition of CS inhibited HUVEC vessel formation, other factors may be present in NCCM and NCM that affect neurite and vessel growth. Therefore, future studies testing an NC-based regenerative strategy should carefully assess the risk of such adverse effects in an in vivo setting