Is quercetin an alternative natural crosslinking agent to genipin for long‐term dermal scaffolds implantation?

As biocompatible matrices, porcine dermal scaffolds have limited application in tissue engineering due to rapid degradation following implantation. This study compared the physical, chemical and biomechanical changes that occurred when genipin and quercetin were used to crosslink dermal scaffolds and to determine whether quercetin could be used as an alternative to genipin. Physicochemical changes in the collagen were assessed using spectroscopic methods [X‐ray diffraction analysis (XRD) and nuclear magnetic resonance (NMR) analysis]. The crosslinking reaction was evaluated by quantification of amino acids and the degree of this reaction by ninhydrin assay. Because the mechanical behaviour of the collagen matrices is highly influenced by crosslinking, the tensile strength of both sets of scaffolds was evaluated. The highest mechanical strength, stiffness, degree of crosslinking and changes in the packing features of collagen (measured by XRD) were achieved using genipin. Some of the results found in the quercetin‐crosslinked scaffolds were possibly due to hydration and dehydration effects elicited by the solvents (phosphate‐buffered saline or ethanol), as seen in the NMR results. In the quercetin‐ethanol‐crosslinked scaffolds, possible reorientation of the amino groups of the collagen molecule may have taken place. Therefore, depending on their proximity to the crosslinking reagent, different types and numbers of interactions may have occurred, inducing a higher crosslinking degree (as evidenced by the ninhydrin assay) and reduction in the free amino acids after reaction. Both crosslinking agents and solvents interfere in the physicochemical properties of collagen thereby inducing variations in the matrix structure. Quercetin‐crosslinked scaffolds may have broader clinical application where a lower degree of crosslinking and stiffness is required. Copyright © 2016 John Wiley & Sons, Ltd.     

An Observational Study of Paediatric Preoperative Transfusion Practice in a Resource-Limited Setting

World Journal of Surgery - Paediatric anaemia is highly prevalent in low–middle-income countries and can negatively impact postoperative outcomes. Currently, there are no guidelines for the...     

PCR can help early diagnosis of pulmonary tuberculosis

One hundred and fifty-one patients, clinically suspected for pulmonary tuberculosis (age: 31 +/- 13 years, male/female: 112/39), were investigated to evaluate the diagnostic potential of polymerase chain reaction (PCR) based detection of the Mycobacterium tuberculosis complex in sputum. The diagnostic efficacy of PCR was compared with culture of Mycobacterium tuberculosis on egg-based Lowenstein-Jensen modified medium. PCR detected 71.5% (108/151), whereas culture detected 66.2% (100/151) of the clinically suspected patients. There was a significant association between the results of PCR and culture (chi2 = 59.524, p < 0.001). However, 23.2% (35/151) samples were found negative in both culture and PCR. Considering culture as the gold standard, the sensitivity of the PCR was 92%. and its specificity 70%. This lower apparent specificity may be due to the higher sensitivity of PCR.     

Protection against acoustic trauma by forward and backward sound conditioning

The purpose of the present study was to determine if short-term sound conditioning provides protection when delivered either before (forward sound conditioning) or after (backward sound conditioning) a traumatic exposure in the guinea pig. Two different sound conditioning paradigms were studied (1 kHz, 81 dB SPL, 24 h; 6.3 kHz, 78 dB SPL, 24 h). The 1-kHz forward sound conditioning paradigm (81 dB SPL, 24 h) protected distortion product otoacoustic emissions (DPOAEs) against a short-duration acoustic trauma (2.7 kHz, 103 dB SPL, 5 min) compared to the group exposed to the acoustic trauma alone. The 1-kHz forward sound conditioning paradigm (81 dB SPL, 24 h) also protected both the auditory brainstem response (ABR) thresholds and DPOAEs against a longer-duration acoustic trauma (2.7 kHz, 103 dB SPL, 30 min). The group exposed to the acoustic trauma alone showed ABR threshold shifts between 15 and 24 dB, and DPOAE amplitude shifts between 11 and 24 dB, while the group with 1-kHz forward sound conditioning showed statistically significant protection at all ABR frequencies and at all DPOAE frequencies. The 1-kHz backward sound conditioning paradigm protected against acoustic trauma (2.7 kHz, 103 dB SPL, 30 min). The ABR thresholds were protected at 1, 2 and 4 kHz, and DPOAEs at all frequencies (except 8 kHz) when compared to the group exposed only to the acoustic trauma. The 6.3-kHz forward sound conditioning paradigm protected against acoustic trauma (5.5 kHz, 109 dB SPL, 30 min) at 6.3, 8 and 10 kHz. The 6.3-kHz backward sound conditioning paradigm showed no protection against acoustic trauma at any DPOAE frequency. Taken together, these findings are important for understanding how the auditory system can be modulated by acoustic stimulation and highlights the importance of the acoustic environment during the recovery process of the auditory system.     

Development of a visual inspection programme for cervical cancer prevention in Bangladesh

Cervical cancer is the most common reproductive cancer in women in Bangladesh, and most women come for diagnosis and treatment when it is too late. To support early detection of pre-cancerous conditions and prevent cervical cancer, Bangladesh undertook a screening programme using visual inspection of the cervix with acetic acid (VIA) and cryotherapy through a public sector programme. The programme was launched in 2004−05 in a phased manner, starting with a pilot programme in 16 of the 64 districts in the country and scaled up to 44 districts as of the end of 2007. Evaluation of the pilot programme's performance showed that VIA can be carried out by trained doctors, nurses, and paramedical workers in Bangladesh, even though the level of resources is poor, and women, their partners and families are often not aware of the disease and its consequences. The programme now needs to move from opportunistic screening to population-based, systematic screening of women over age 30. More providers need to be trained, and clinics better equipped. The links between screening, diagnosis and treatment need to be improved and the false-positive rate of VIA tests greatly reduced. It is only when we have achieved high coverage that reduced rates of cancer can be ensured.     

The development and implantation of a biologically derived allograft scaffold

Biologically derived scaffolds are becoming viable treatment options for tissue/organ repair and regeneration. A continuing hurdle is the need for a functional blood supply to and from the implanted scaffold. We have addressed this problem by constructing an acellular ileal scaffold with an attached vascular network suitable for implantation and immediate reperfusion with the host's blood. Using a vascular perfusion approach, a segment of porcine ileum up to 30 cm long, together with its attached vasculature, was decellularized as a single entity. The quality of the decellularized scaffold was assessed histologically and using molecular tools. To establish vascular perfusion potentials of the scaffold, a right‐sided nephrectomy and end‐to‐end anastomosis of the decellularized scaffold's vasculature to a renal artery and vein were performed in a pig of similar size to the donor animal. Lengths of ileal scaffold, together with its attached vasculature, were successfully decellularized, with no evidence of intact cells/nuclear material or collagen degradation. The scaffold's decellularized vascular network demonstrated optimum perfusion at 1, 2 and 24 h post‐implantation and the mesenteric arcade remained patent throughout the assessment. The 1, 2 and 24 h explanted scaffolds demonstrated signs of cellular attachment, with cells positive for CD68 and CD133 on the vascular luminal aspect. It is possible to decellularize clinically relevant lengths of small intestine, together with the associated vasculature, as a single segment. The functional vascular network may represent a route for recellularization for future regeneration of bowel tissue for patients with short bowel syndrome. Copyright © 2013 John Wiley & Sons, Ltd.     

p19ARF is a critical mediator of both cellular senescence and an innate immune response associated with MYC inactivation in mouse model of acute leukemia

MYC-induced T-ALL exhibit oncogene addiction. Addiction to MYC is a consequence of both cell-autonomous mechanisms, such as proliferative arrest, cellular senescence, and apoptosis, as well as non-cell autonomous mechanisms, such as shutdown of angiogenesis, and recruitment of immune effectors. Here, we show, using transgenic mouse models of MYC-induced T-ALL, that the loss of either p19ARF or p53 abrogates the ability of MYC inactivation to induce sustained tumor regression. Loss of p53 or p19ARF, influenced the ability of MYC inactivation to elicit the shutdown of angiogenesis; however the loss of p19ARF, but not p53, impeded cellular senescence, as measured by SA-beta-galactosidase staining, increased expression of p16INK4A, and specific histone modifications. Moreover, comparative gene expression analysis suggested that a multitude of genes involved in the innate immune response were expressed in p19ARF wild-type, but not null, tumors upon MYC inactivation. Indeed, the loss of p19ARF, but not p53, impeded the in situ recruitment of macrophages to the tumor microenvironment. Finally, p19ARF null-associated gene signature prognosticated relapse-free survival in human patients with ALL. Therefore, p19ARF appears to be important to regulating cellular senescence and innate immune response that may contribute to the therapeutic response of ALL.     

Immunomodulatory effect of a decellularized skeletal muscle scaffold in a discordant xenotransplantation model

Decellularized (acellular) scaffolds, composed of natural extracellular matrix, form the basis of an emerging generation of tissue-engineered organ and tissue replacements capable of transforming healthcare. Prime requirements for allogeneic, or xenogeneic, decellularized scaffolds are biocompatibility and absence of rejection. The humoral immune response to decellularized scaffolds has been well documented, but there is a lack of data on the cell-mediated immune response toward them in vitro and in vivo. Skeletal muscle scaffolds were decellularized, characterized in vitro, and xenotransplanted. The cellular immune response toward scaffolds was evaluated by immunohistochemistry and quantified stereologically. T-cell proliferation and cytokines, as assessed by flow cytometry using carboxy-fluorescein diacetate succinimidyl ester dye and cytometric bead array, formed an in vitro surrogate marker and correlate of the in vivo host immune response toward the scaffold. Decellularized scaffolds were free of major histocompatibility complex class I and II antigens and were found to exert anti-inflammatory and immunosuppressive effects, as evidenced by delayed biodegradation time in vivo; reduced sensitized T-cell proliferative activity in vitro; reduced IL-2, IFN-γ, and raised IL-10 levels in cell-culture supernatants; polarization of the macrophage response in vivo toward an M2 phenotype; and improved survival of donor-derived xenogeneic cells at 2 and 4 wk in vivo. Decellularized scaffolds polarize host responses away from a classical TH1-proinflammatory profile and appear to down-regulate T-cell xeno responses and TH1 effector function by inducing a state of peripheral T-cell hyporesponsiveness. These results have substantial implications for the future clinical application of tissue-engineered therapies.Although replacement of airways (1–6) or urogenital tissue (7) using stem cell-based techniques has been achieved, engineering of functional contractile muscle tissue has only been partially explored (8–9). While debates have been generated around which myogenic progenitors (satellite cells, muscle stem cells, or myoblasts) should be used, fewer studies have focused on exploring which matrix could offer a better platform for regeneration (10, 11).Autologous tissue-engineered solutions have the major advantage of not requiring immunosuppression, but clinical applications are limited to static organs and tissues, such as skin, or those that can function through passive movement alone, such as trachea, heart valves, blood vessels, and bladder (4, 7, 12). The field continues to expand and tissue bioengineering has provided, or is close to delivering, functional human organ replacements elsewhere (6, 7, 13–17). The ability to produce innervated and revascularized muscles would hugely extend the possible applications of regenerative medicine (18–26).Decellularized skeletal muscle has been characterized by several groups, but its effect on cell-mediated immunity has not been studied (27–30). Here, we provide evidence that decellularized muscle scaffolds promote anti-inflammatory and immunosuppressive responses both in vitro and in vivo, down-regulate T-cell xeno responses and TH1 effector cytokines in vitro, and polarize the macrophage response in vivo toward an M2 phenotype (i.e., promote alternative pathway activation of macrophages).