Redifferentiated cardiomyocytes retain residual dedifferentiation signatures and are protected against ischemic injury.

Cardiomyocyte proliferation and dedifferentiation have fueled the field of regenerative cardiology in recent years, whereas the reverse process of redifferentiation remains largely unexplored. Redifferentiation is characterized by the restoration of function lost during dedifferentiation. Previously, we showed that ERBB2-mediated heart regeneration has these two distinct phases: transient dedifferentiation and redifferentiation. Here we survey the temporal transcriptomic and proteomic landscape of dedifferentiation-redifferentiation in adult mouse hearts and reveal that well-characterized dedifferentiation features largely return to normal, although elements of residual dedifferentiation remain, even after the contractile function is restored. These hearts appear rejuvenated and show robust resistance to ischemic injury, even 5 months after redifferentiation initiation. Cardiomyocyte redifferentiation is driven by negative feedback signaling and requires LATS1/2 Hippo pathway activity. Our data reveal the importance of cardiomyocyte redifferentiation in functional restoration during regeneration but also protection against future insult, in what could lead to a potential prophylactic treatment against ischemic heart disease for at-risk patients.

Tracheal In Vitro Reconstruction Using a Decellularized Bio-Scaffold in Combination with a Rotating Bioreactor.

Long-segment airway stenosis as well as their neoplastic transformation is life-threatening and still currently represent unsolved clinical problems. Indeed, despite several attempts, definitive surgical procedures are not presently available, and a suitable tracheal reconstruction or replacement remains an urgent clinical need. A possible innovative strategic solution to restore upper airway function may be represented by the creation of a bioprosthetic trachea, obtained through the combination of tissue engineering and regenerative medicine.Here we describe a two-step protocol for the ex vivo generation of tracheal segments. The first step involves the application of a decellularization technique that allows for the production of a naturally derived extracellular matrix (ECM)-based bio-scaffold, that maintains the macro- and micro-architecture as well as 9 the matrix-related signals distinctive of the original tissue. In the second step chondrocytes are seeded onto decellularized trachea, using a rotating bioreactor to ensure a correct scaffold repopulation.This multi-step approach represents a powerful tool for in vitro reconstruction of a bioengineered trachea that may constitute a promising solution to restore upper airway function. In addition, the procedures here described allow for the creation of a suitable 3D platform that may find useful applications, both for toxicological studies as well as organ transplantation strategies.

Literature Review and Knowledge Distribution During an Outbreak: A Methodology for Managing Infodemics.

PROBLEM: The COVID-19 pandemic has challenged health care systems in an unprecedented way by imposing new demands on health care resources and scientific knowledge. There has also been an exceedingly fast accumulation of new information on this novel virus. As the traditional peer-review process takes time, there is currently a significant gap between the ability to generate new data and the ability to critically evaluate them. This problem of an excess of mixed-quality data, or infodemic, is echoing throughout the scientific community. APPROACH: The authors aimed to help their colleagues at the Rambam Medical Center, Haifa, Israel, manage the COVID-19 infodemic with a methodologic solution: establishing an in-house mechanism for continuous literature review and knowledge distribution (March-April 2020). Their methodology included the following building blocks: a dedicated literature review team, artificial intelligence-based research algorithms, brief written updates in a graphical format, large-scale webinars and online meetings, and a feedback loop. OUTCOMES: During the first month (April 2020), the project produced 21 graphical updates. After consideration of feedback from colleagues and final editing, 13 graphical updates were uploaded to the center's website; of these, 31% addressed the clinical presentation of the disease and 38% referred to specific treatments. This methodology as well as the graphical updates it generated were adopted by the Israeli Ministry of Health and distributed in a hospital preparation kit. NEXT STEPS: The authors believe they have established a novel methodology that can assist in the battle against COVID-19 by making high-quality scientific data more accessible to clinicians. In the future, they expect this methodology to create a favorable uniform standard for evidence-guided health care during infodemics. Further evolution of the methodology may include evaluation of its long-term sustainability and impact on the day-to-day clinical practice and self-confidence of clinicians who treat COVID-19 patients.

Using Decellularization/Recellularization Processes to Prepare Liver and Cardiac Engineered Tissues.

Tissue engineering provides unique opportunities for disease modeling, drug testing, and regenerative medicine applications. The use of cell-seeded scaffolds to promote tissue development is the hallmark of the tissue engineering. Among the different types of scaffolds (derived from either natural or synthetic polymers) used in the field, the use of decellularized tissues/organs is specifically attractive. The decellularization process involves the removal of native cells from the original tissue, allowing for the preservation of the three-dimensional (3D) macroscopic and microscopic structures of the tissue and extracellular matrix (ECM) composition. Following recellularization, the resulting scaffold provides the seeded cells with the appropriate biological signals and mechanical properties of the original tissue. Here, we describe different methods to create viable scaffolds from decellularized heart and liver as useful tools to study and exploit ECM biological key factors for the generation of engineered tissues with enhanced regenerative properties.

Creation of a Bioengineered Ovary: Isolation of Female Germline Stem Cells for the Repopulation of a Decellularized Ovarian Bioscaffold.

Ovarian failure is the most common cause of infertility and affects about 1% of young women. One innovative strategy to restore ovarian function may be represented by the development of a bioprosthetic ovary, obtained through the combination of tissue engineering and regenerative medicine.We here describe the two main steps required for bioengineering the ovary and for its ex vivo functional reassembling. The first step aims at producing a 3D bioscaffold, which mimics the natural ovarian milieu in vitro. This is obtained with a whole organ decellularization technique that allows the maintenance of microarchitecture and biological signals of the original tissue. The second step involves the use of magnetic activated cell sorting (MACS) to isolate purified female germline stem cells (FGSCs). These cells are able to differentiate in ovarian adult mature cells, when subjected to specific stimuli, and can be used them to repopulate ovarian decellularized bioscaffolds. The combination of the two techniques represents a powerful tool for in vitro recreation of a bioengineered ovary that may constitute a promising solution for hormone and fertility function restoring. In addition, the procedures here described allow for the creation of a suitable 3D platform with useful applications both in toxicological and transplantation studies.

Osteopathic Cranial Manipulation for a Patient With Whiplash-Associated Disorder: A Case Report.

Objective: The purpose of this case report is to describe the immediate effects of osteopathic cranial manipulation on pain and cervical motion in a patient with whiplash-associated disorder. Clinical Features: A 74-year-old man reported daily cervicogenic headaches after a whiplash injury caused by a traffic accident 3 months prior. Physical examination and osteopathic assessment identified tissue texture alteration, positional asymmetry, limited range of motion, and tenderness. The resulting diagnosis was somatic dysfunction of the head and the cervical region. Intervention and Outcome: Osteopathic manipulative treatment was administered to the occipital area. The patient reported an immediate improvement in pain. Accessory movement of the cervical spine was improved. Conclusion: This patient responded favorable to osteopathic cranial manipulation with improved symptoms and ranges of motion.

Bioengineering the ovary to preserve and reestablish female fertility.

Different bioengineering strategies can be presently adopted and have been shown to have great potential in the treatment of female infertility and ovarian dysfunction deriving from chemotherapy, congenital malformations, massive adhesions as well as aging and lifestyle. One option is transplantation of fresh or cryopreserved organs/fragments into the patient. A further possibility uses tissue engineering approaches that involve a combination of cells, biomaterials and factors that stimulate local ability to regenerate/ repair the reproductive organ. Organ transplant has shown promising results in large animal models. However, the source of the organ needs to be identified and the immunogenic effects of allografts remain still to be solved before the technology may enter the clinical practice. Decellularization/ repopulation of ovary with autologous cells or follicles could represent an interesting, still very experimental alternative. Here we summarize the recent advancements in the bioengineering strategies applied to the ovary, we present the principles for these systems and discuss the advantages of these emerging opportunities to preserve or improve female fertility.

Whole-ovary decellularization generates an effective 3D bioscaffold for ovarian bioengineering.

PURPOSE: To develop a new protocol for whole-ovary decellularization for the production of a 3D bioscaffold suitable for in vitro/ex vivo studies and for the reconstruction of a bioengineered ovary. METHODS: Porcine ovaries were subjected to the decellularization process (DECELL; n = 20) that involved a freeze-thaw cycle, followed by sequential incubations in 0.5% SDS for 3 h, 1% Triton X-100 for 9 h, and 2% deoxycholate for 12 h. Untreated ovaries were used as a control (CTR; n = 6). Both groups were analyzed to evaluate cell and DNA removal as well as ECM preservation. DECELL bioscaffolds were assessed for cytotoxicity and cell homing ability. RESULTS: DECELL ovaries maintained shape and homogeneity without any deformation, while their color turned from red to white. Histological staining and DNA quantification confirmed a decrease of 98.11% in DNA content, compared with the native tissue (CTR). Histochemical assessments demonstrated the preservation of intact ECM microarchitecture after the decellularization process. This was also confirmed by quantitative analysis of collagen, elastin, and GAG contents. DECELL bioscaffold showed no cytotoxic effects in co-culture and, when re-seeded with homologous fibroblasts, encouraged a rapid cell adhesion and migration, with repopulating cells increasing in number and aggregating in cluster-like structures, consistent with its ability to sustain cell adherence, proliferation, and differentiation. CONCLUSION: The protocol described allows for the generation of a 3D bioscaffold that may constitute a suitable model for ex vivo culture of ovarian cells and follicles, as well as a promising tool for the reconstruction of a bioengineered ovary.

Simple and Quick Method to Obtain a Decellularized, Functional Liver Bioscaffold.

The development of new approaches for organ transplantation has become crucial in the last years. In particular, organ engineering, involving the preparation of acellular matrices that provide a natural habitat for reseeding with an appropriate population of cells, is an attractive although technically demanding approach. We here describe a method that allows for the derivation of functional in vitro hepatic organoids and that does not require a previous selection of all the parenchymal hepatocytes and non-parenchymal cells, namely, Kupffer cells, liver endothelial cells, and hepatic stellate cells. The procedure also replaces the costly standard collagenase perfusion step with a trypsin-based enzymatic digestion that results in high-yield decellularization. A combination of physical and chemical treatments through deep immersion and intraluminal infusion of two different consecutive solutions is used: (1) deionized water (DI) and (2) DI + Triton X 1% + ammonium hydroxide (NH4OH) 0.1%. This ensures the isolation of the hepatic constructs that reliably maintain original architecture and ECM components while completely removing cellular DNA and RNA. The procedure is fast, simple, and cheap and warrants an optimal organoid functionality that may find applications in both toxicological and transplantation studies.

High Penetration of Paclitaxel in Abdominal Wall of Rabbits after Hyperthermic Intraperitoneal Administration of Nab-Paclitaxel Compared to Standard Paclitaxel Formulation.

PURPOSE: Paclitaxel (PTX) is currently used in combination with cisplatin for Hyperthermic Intraperitoneal Chemotherapy (HIPEC) for the treatment of peritoneal carcinomatosis. Albumin-bound PTX is a promising new drug for HIPEC because of its easy solubility in aqueous perfusion medium and possibly because of the tendency of albumin to cross physiological barriers and accumulate in tumor tissue. METHODS: We tested the feasibility of using nab-paclitaxel in rabbits treated by HIPEC for 60 min compared with the classical formulation at an equivalent PTX dose. Samples of perfusate and blood were collected at different time points and peritoneal tissues were collected at the end of perfusion. PTX concentrations were determined by HPLC. The depth of paclitaxel penetration through the peritoneal barrier was assessed by mass spectrometry imaging. RESULTS: PTX after nab-paclitaxel treatment penetrated up to 0.63 mm in the peritoneal wall, but after CRE-paclitaxel, it was not detectable in the peritoneum. Moreover, the peritoneal concentration after nab-paclitaxel was five times that after paclitaxel classical formulation. Despite the high levels reached in the peritoneum, systemic exposure of PTX was low. CONCLUSIONS: Our results show that nab-paclitaxel penetrates into the abdominal wall better than CRE-paclitaxel, in terms of effective penetration and peritoneal tissue concentration.