Application of pulsed electric field technology to skin engineering.

Tissue engineering encompasses a range of techniques that direct the growth of cells into a living tissue construct for regenerative medicine applications, disease models, drug discovery, and safety testing. These techniques have been implemented to alleviate the clinical burdens of impaired healing of skin, bone, and other tissues. Construct development requires the integration of tissue-specific cells and/or an extracellular matrix-mimicking biomaterial for structural support. Production of such constructs is generally expensive and environmentally costly, thus eco-sustainable approaches should be explored. Pulsed electric field (PEF) technology is a nonthermal physical processing method commonly used in food production and biomedical applications. In this review, the key principles of PEF and the application of PEF technology for skin engineering will be discussed, with an emphasis on how PEF can be applied to skin cells to modify their behaviour, and to biomaterials to assist in their isolation or sterilisation, or to modify their physical properties. The findings indicate that the success of PEF in tissue engineering will be reliant on systematic evaluation of key parameters, such as electric field strength, and their impact on different skin cell and biomaterial types. Linking tangible input parameters to biological responses critical to healing will assist with the development of PEF as a sustainable tool for skin repair and other tissue engineering applications.

Understanding the effect of pulsed electric fields on thermostability of connective tissue isolated from beef pectoralis muscle using a model system.

Brisket is a low value/tough meat cut that contains a large amount of connective tissue. Conversion of collagen into gelatin during heating reduces the toughness of the connective tissue however this conversion is slow at low cooking temperatures (around 60°C). The objective of this project was to determine the ability of pulsed electric field (PEF) processing to reduce the thermal stability of connective tissue. To achieve this, a novel model system was designed in which connective tissue obtained from beef deep pectotalis muscle (brisket) was exposed to PEF at combinations of electric field strength (1.0 and 1.5kV/cm) and specific energy (50 and 100kJ/kg) within an agar matrix at electrical conductivities representing the electrical conductivity found in brisket. Differential scanning calorimetry showed that PEF treatment significantly (p<0.05) decreased the denaturation temperature of connective tissue compared to untreated samples. Increasing electric field strength and the specific energy increased the Ringer soluble collagen fraction. PEF treated samples showed higher solubilization compared to the untreated samples at both 60°C and 70°C in heat solubility test. SEM examination of PEF treated (at 1.5kV/cm and 100kJ/kg) and untreated samples revealed that PEF appeared to increase the porosity of the connective tissue structure. These finding suggest that PEF processing is a technology that could be used to improve the tenderness and decrease the cooking time of collagen rich, meat cuts.

Critical analysis of the maximum non inhibitory concentration (MNIC) method in quantifying sub-lethal injury in Saccharomyces cerevisiae cells exposed to either thermal or pulsed electric field treatments.

Sub-lethal injury within a microbial population, due to processing treatments or environmental stress, is often assessed as the difference in the number of cells recovered on non-selective media compared to numbers recovered on a "selective media" containing a predetermined maximum non-inhibitory concentration (MNIC) of a selective agent. However, as knowledge of cell metabolic response to injury, population diversity and dynamics increased, the rationale behind the conventional approach of quantifying sub-lethal injury must be scrutinized further. This study reassessed the methodology used to quantify sub-lethal injury for Saccharomyces cerevisiae cells (≈ 4.75 Log CFU/mL) exposed to either a mild thermal (45°C for 0, 10 and 20min) or a mild pulsed electric field treatment (field strengths of 8.0-9.0kV/cm and energy levels of 8, 14 and 21kJ/kg). Treated cells were plated onto either Yeast Malt agar (YM) or YM containing NaCl, as a selective agent at 5-15% in 1% increments. The impact of sub-lethal stress due to initial processing, the stress due to selective agents in the plating media, and the subsequent variation of inhibition following the treatments was assessed based on the CFU count (cell numbers). ANOVA and a generalised least squares model indicated significant effects of media, treatments, and their interaction effects (P<0.05) on cell numbers. It was shown that the concentration of the selective agent used dictated the extent of sub-lethal injury recorded owing to the interaction effects of the selective component (NaCl) in the recovery media. Our findings highlight a potential common misunderstanding on how culture conditions impact on sub-lethal injury. Interestingly for S. cerevisiae cells the number of cells recovered at different NaCl concentrations in the media appears to provide valuable information about the mode of injury, the comparative efficacy of different processing regimes and the inherent degree of resistance within a population. This approach may provide similar information for other micro-organisms.

L-ascorbic acid improves the serum folate response to an oral dose of [6S]-5-methyltetrahydrofolic acid in healthy men.

OBJECTIVE: To investigate the effect of simultaneous administration of [6S]-5-methyltetrahydrofolic acid ([6S]-5-CH(3)H(4)PteGlu) with L-ascorbic acid (L-AA) on serum folate concentrations in healthy male subjects. SUBJECTS AND METHODS: A total of nine healthy male volunteers were recruited. Serum folate concentrations were measured before and up to 8 h after administration of each treatment (1) placebo, (2) 343 microg [6S]-5-CH(3)H(4)PteGlu), (3) 343 microg [6S]-5-CH(3)H(4)PteGlu) with 289.4 mg L-AA and (4) 343 microg [6S]-5-CH(3)H(4)PteGlu) with 973.8 mg L-AA (n=10 samples per treatment). RESULTS: Serum folate concentrations significantly increased compared with baseline values, starting from 30 min after [6S]-5-CH(3)H(4)PteGlu administration and remained significantly higher than baseline values during the first 6 h for treatments 3 and 4, and during the first 4 h for treatment 2. Maximal serum folate responses were observed between 0.5 and 1.5 h after [6S]-5-CH(3)H(4)PteGlu consumption and significantly differed between treatments 2 and 4 (P<0.05). When [6S]-5-CH(3)H(4)PteGlu was concurrently administered with 289.4 or 973.8 mg L-AA, the total serum folate response, calculated as the area under the curve (AUC), was significantly improved (46.5+/-4.0 and 53.0+/-4.0 vs 34.3+/-3.8 h nmol/l, P<0.05). No significant difference in AUC was found between the 289.4 and the 973.8 mg L-AA treatments. CONCLUSIONS: Administration of a physiological dose of [6S]-5-CH(3)H(4)PteGlu with L-AA significantly improved the measured serum folate response in folate saturated healthy men.

Kinetics of the stability of broccoli (Brassica oleracea Cv. Italica) myrosinase and isothiocyanates in broccoli juice during pressure/temperature treatments.

The Brassicaceae plant family contains high concentrations of glucosinolates, which can be hydrolyzed by myrosinase yielding products having an anticarcinogenic activity. The pressure and temperature stabilities of endogenous broccoli myrosinase, as well as of the synthetic isothiocyanates sulforaphane and phenylethyl isothiocyanate, were studied in broccoli juice on a kinetic basis. At atmospheric pressure, kinetics of thermal (45-60 degrees C) myrosinase inactivation could be described by a consecutive step model. In contrast, only one phase of myrosinase inactivation was observed at elevated pressure (100-600 MPa) combined with temperatures from 10 up to 60 degrees C, indicating inactivation according to first-order kinetics. An antagonistic effect of pressure (up to 200 MPa) on thermal inactivation (50 degrees C and above) of myrosinase was observed indicating that pressure retarded the thermal inactivation. The kinetic parameters of myrosinase inactivation were described as inactivation rate constants (k values), activation energy (Ea values), and activation volume (Va values). On the basis of the kinetic data, a mathematical model describing the pressure and temperature dependence of myrosinase inactivation rate constants was constructed. The stability of isothiocyanates was studied at atmospheric pressure in the temperature range from 60 to 90 degrees C and at elevated pressures in the combined pressure-temperature range from 600 to 800 MPa and from 30 to 60 degrees C. It was found that isothiocyanates were relatively thermolabile and pressure stable. The kinetics of HP/T isothiocyanate degradation could be adequately described by a first-order kinetic model. The obtained kinetic information can be used for process evaluation and optimization to increase the health effect of Brassicaceae.

Postoperative pain detracts from early health status improvement seen after video-assisted thoracoscopic lung volume reduction surgery.

OBJECTIVES: To assess the impact of lung volume reduction surgery (LVRS) on postoperative pain. METHODS: Fifty-two patients, 34 male/18 female, median age 59 (46-70) years, underwent unilateral video-assisted thoracoscopic (VAT) LVRS. FEV(1), TLC, RV and RV/TLC ratio were assessed preoperatively and at 3, 6, 12 and 24 months post surgery. At the same time interval health status was assessed by Euroquol and SF 36 questionnaires. RESULTS: Significant improvements in health status, as assessed by SF 36, persisted from 3 months to 1 year. However, in the pain domain there was a worsening of the mean score from 74 preoperatively to 64 at 3 months, 68 at 6 months, 73 at 12 months and 65 at 24 months. The improvements in Euroquol score were not statistically significant. However, they became significant for at least 2 years postoperatively, when those patients who had a worsening pain score postoperatively were excluded. While the percentage of patients with a worsening of pain scores measured with SF 36 remained between 40 and 45% even 2 years after LVRS, when using Euroquol this percentage did decrease from 30% at 3 months to 14% at 2 years. There was no significant correlation between the change of scores and length of operation, hospital stay or air leak. It was also not statistically significant whether these patients had an extra procedure (redo thoracotomy or insertion of extra drain postoperatively). There were some significant correlations between changes in hyperinflation and changes in pain scores but this was not consistent for Euroquol and SF 36. CONCLUSION: Postoperative pain detracts from global improvement in health status after LVRS even after unilateral VATS. There may be an influence of alterations in chest mechanics after surgery on the development of pain.

En-bloc chest wall and lung resection for non-small cell lung cancer. Predictors of 60-day non-cancer related mortality.

OBJECTIVE: Predictors for early mortality after isolated pulmonary resection have been identified and clear guidelines regarding fitness for surgery have been produced. However, the additional risk of en-bloc chest wall resection has not been extensively studied. AIM: We analyzed our total experience of combined chest wall and lung resection for non-small-cell lung cancer (NSCLC) to identify additional risk factors for early non-tumour related mortality. PATIENTS AND METHODS: A retrospective review of 41 consecutive patients, with median age of 69 (range 37-84) years, operated by a single surgeon over a 4-year period. Univariate analysis was performed to assess the relationship of selected preoperative and operative variables on mortality within 2 months from surgery. RESULTS: Low preoperative body mass index, age over 75 years, and preoperative FEV(1) of less than 70% of predicted were associated with a significantly increased 60-day mortality. In those patients with any of these risk factors 60-day mortality was 47% (8 of 17). In those with none of the above there was no mortality (of 24 patients) (P=0.0004). DISCUSSION: En-bloc pulmonary and chest wall resection for NSCLC should be avoided in the elderly, those with limited respiratory reserve or significant weight loss. These factors render the patient highly susceptible to chest complications leading to increased mortality.

Lung volume reduction surgery--a comparison of the long term outcome of unilateral vs. bilateral approaches.

OBJECTIVE: Bilateral lung volume reduction surgery (LVRS) is thought to be preferable to unilateral surgery due to greater initial benefit but the subsequent rate of decline may also be greater. We compared the long term physiological and health status outcome of LVRS performed on one or simultaneously on both lungs. METHODS: Prospective data were collected on a consecutive series of 65 patients undergoing LVRS who were all suitable for bilateral surgery. Twenty-six patients: age 59 (8) years underwent bilateral LVRS by video-assisted thoracoscopy (VAT) or sternotomy and 39 patients: age 60 (6) years underwent unilateral VAT. The perioperative effects of LVRS on spirometry were prospectively recorded at 3, 6, 12 and 24 months. RESULTS: The unilateral group had similar preoperative lung volumes to the bilateral patients: forced expiratory volume in 1s (FEV(1)) 26 vs. 30% predicted, RV 275 vs. 246% predicted and total lung capacity (TLC) 148 vs. 142% predicted. Unilateral LVRS was associated with significantly lower weight of lung resected: 80 (31) vs. 118 (46) g; hospital stay: 16 (10) days vs. 28 (22) days. Thirty-day mortality was 3% in the unilateral and 8% in the bilateral group (P=0.34). Postoperative ventilation occurred in 5% in the unilateral and in 42% in the bilateral group (P=0.0002). The decline of FEV(1) during the first postoperative year was significant in the bilateral group (-313 ml/y, P=0.04) but not significant in the unilateral group (-50 ml/y, P=0.18). SF 36 scores in all eight domains were similar in both groups preoperatively and at any postoperative interval. CONCLUSION: We have found no benefit from bilateral simultaneous LVRS and prefer unilateral LVRS because of the lower morbidity, resulting in earlier discharge, and slower decline in physiological benefit.

Post-pneumonectomy video-assisted thoracoscopic bullectomy using extra-corporeal membrane oxygenation.

We describe a case of a patient who, 14 years after a pneumonectomy, required surgery for a life-threatening air-leak following accidental intubation of an emphysematous bulla in his remaining lung. To facilitate treatment by video-assisted thoracoscopic surgery, veno-venous extra-corporeal membrane oxygenation was employed.

Metalloptysis: a late complication of lung volume reduction surgery.

We describe three cases where patients expectorated titanium staples many months after lung volume reduction surgery (LVRS). The possible mechanisms and technical implications of this rare complication are discussed.