Christopher Glenn Wallace

Melatonin treatment improves adipose-derived mesenchymal stem cell therapy for acute lung ischemia–reperfusion injury

This study investigated whether melatonin‐treated adipose‐derived mesenchymal stem cells (ADMSC) offered superior protection against acute lung ischemia–reperfusion (IR) injury. Adult male Sprague‐Dawley rats (n = 30) were randomized equally into five groups: sham controls, lung IR–saline, lung IR–melatonin, lung IR–melatonin–normal ADMSC, and lung IR–melatonin–apoptotic ADMSC. Arterial oxygen saturation was lowest in lung IR–saline; lower in lung IR–melatonin than sham controls, lung IR–melatonin–normal ADMSC, and lung IR–melatonin–apoptotic ADMSC; lower in lung IR–melatonin–normal ADMSC than sham controls and lung IR–melatonin–apoptotic ADMSC; lower in lung IR–melatonin–apoptotic ADMSC than sham controls (P < 0.0001 in each case). Right ventricular systolic blood pressure (RVSBP) showed a reversed pattern among all groups (all P < 0.0001). Changes in histological scoring of lung parenchymal damage and CD68+ cells showed a similar pattern compared with RVSBP in all groups (all P < 0.001). Changes in inflammatory protein expressions such as VCAM‐1, ICAM‐1, oxidative stress, TNF‐α, NF‐κB, PDGF, and angiotensin II receptor, and changes in apoptotic protein expressions of cleaved caspase 3 and PARP, and mitochondrial Bax, displayed identical patterns compared with RVSBP in all groups (all P < 0.001). Numbers of antioxidant (GR+, GPx+, NQO‐1+) and endothelial cell biomarkers (CD31+ and vWF+) were lower in sham controls, lung IR–saline, and lung IR–melatonin than lung IR–melatonin–normal ADMSC and lung IR–melatonin–apoptotic ADMSC, and lower in lung IR–melatonin–normal ADMSC than lung IR–melatonin–apoptotic ADMSC (P < 0.001 in each case). In conclusion, when the animals were treated with melatonin, the apoptotic ADMSC were superior to normal ADMSC for protection of lung from acute IR injury.

Additional benefit of combined therapy with melatonin and apoptotic adipose-derived mesenchymal stem cell against sepsis-induced kidney injury

This study tested whether combined therapy with melatonin and apoptotic adipose‐derived mesenchymal stem cells (A‐ADMSCs) offered additional benefit in ameliorating sepsis‐induced acute kidney injury. Adult male Sprague–Dawley rats (n = 65) were randomized equally into five groups: Sham controls (SC), sepsis induced by cecal‐ligation and puncture (CLP), CLP‐melatonin, CLP‐A‐ADMSC, and CLP‐melatonin‐A‐ADMSC. Circulating TNF‐α level at post‐CLP 6 hr was highest in CLP and lowest in SC groups, higher in CLP‐melatonin than in CLP‐A‐ADMSC and CLP‐melatonin‐A‐ADMSC groups (all P < 0.001). Immune reactivity as reflected in the number of splenic helper‐, cytoxic‐, and regulatory‐T cells at post‐CLP 72 hr exhibited the same pattern as that of circulating TNF‐α among all groups (P < 0.001). The histological scoring of kidney injury and the number of F4/80+ and CD14+ cells in kidney were highest in CLP and lowest in SC groups, higher in CLP‐melatonin than in CLP‐A‐ADMSC and CLP‐melatonin‐A‐ADMSC groups, and higher in CLP‐A‐ADMSC than in CLP‐melatonin‐A‐ADMSC groups (all P < 0.001). Changes in protein expressions of inflammatory (RANTES, TNF‐1α, NF‐κB, MMP‐9, MIP‐1, IL‐1β), apoptotic (cleaved caspase 3 and PARP, mitochondrial Bax), fibrotic (Smad3, TGF‐β) markers, reactive‐oxygen‐species (NOX‐1, NOX‐2), and oxidative stress displayed a pattern identical to that of kidney injury score among the five groups (all P < 0.001). Expressions of antioxidants (GR+, GPx+, HO‐1, NQO‐1+) were lowest in SC group and highest in CLP‐melatonin‐A‐ADMSC group, lower in CLP than in CLP‐melatonin and CLP‐A‐ADMSC groups, and lower in CLP‐melatonin‐ than in CLP‐A‐ADMSC‐tretaed animals (all P < 0.001). In conclusion, combined treatment with melatonin and A‐ADMSC was superior to A‐ADMSC alone in protecting the kidneys from sepsis‐induced injury.

Two small flaps from one anterolateral thigh donor site for bilateral buccal mucosa reconstruction after release of submucous fibrosis and/or contracture

Oral submucous fibrosis causes health-related and social problems for affected patients. Free flap reconstruction has proved effective for maintaining mouth opening after release of fibrosis. Two independent free flaps from separate donor sites, such as bilateral forearm flaps or bilateral anterolateral thigh (ALT) flaps, were traditionally required for reconstruction. The former option sacrifices one of the two major arteries in the forearm. Both options are time consuming and required two donor sites. To eliminate these disadvantages, we developed a technical modification that allows harvesting of two independent flaps from one ALT thigh based on one descending branch of the lateral circumflex femoral artery (d-LCFA). Eighteen flaps from nine donor sites were harvested for post-release reconstruction of oral submucous fibrosis. Mean flap size was 4.1 x 7.5 cm, mean pedicle length was 7.6 cm, mean ischaemia time was 104 min and mean total operation time was 13 h and 19 min. All donor sites were closed primarily, with one exception. One flap failed and was replaced with a contralateral ALT flap. One patient developed a wound infection and another developed a seroma at the recipient site. Four flaps required secondary de-bulking in three patients. The improvement in mouth opening was evaluated by inter-incisor distance (IID): mean preoperative IID was 9.6mm (range: 0-20mm), mean follow-up time was 16.2 months (range: 10-33 months); mean postoperative IID was 23.8mm and mean improvement in IID was 15.3mm (range: 10-27 mm). In conclusion, two independent flaps can be harvested from d-LCFA of the same thigh, instead of from both thighs, to reconstruct bilateral buccal defects after release of submucous fibrosis and/or contracture.

Clinical evidence of field cancerization in patients with oral cavity cancer in a betel quid chewing area

OBJECTIVES We sought to investigate whether there is evidence of field cancerization in patients with oral cavity squamous cell carcinoma (OSCC) enrolled in a betel quid chewing area. We also assessed whether betel quid chewing is an independent risk factor for field cancerization in OSCC patients. METHODS We retrospectively examined the records of 1570 OSCC patients who underwent radical tumor resection between 1996 and 2011. A total of 1243 study participants (79%) had a positive history of betel quid chewing before surgery. Of the 767 patients treated with surgery alone, 599 (78%) were preoperative chewers, whereas a history of preoperative betel quid chewing was identified in 644 (80%) of the 803 patients who received adjuvant therapy. The 5-year control, survival, and second primary tumors (SPTs) rates served as the main outcome measures. RESULTS Regardless of the treatment modality, more than 70% of the SPTs were located in the oral cavity or soft palate. Despite a similar risk profile in terms of tumor depth, lymph node metastasis, and pathological margin status, preoperative chewers showed a significantly higher incidence of 5-year SPTs and local recurrences compared with non-chewers. Moreover, multivariate analysis demonstrated that preoperative betel quid chewing was an independent prognostic factor for 5-year local control and SPTs occurrence rates. CONCLUSIONS Our results demonstrate that preoperative betel quid chewers had a higher incidence of local recurrence and SPTs than non-chewers, suggesting that field cancerization may occur in OSCC patients with a history of betel quid chewing.

Intravenous administration of xenogenic adipose-derived mesenchymal stem cells (ADMSC) and ADMSC-derived exosomes markedly reduced brain infarct volume and preserved neurological function in rat after acute ischemic stroke

We tested the hypothesis that combined xenogenic (from mini-pig) adipose-derived mesenchymal stem cell (ADMSC) and ADMSC-derived exosome therapy could reduce brain-infarct zone (BIZ) and enhance neurological recovery in rat after acute ischemic stroke (AIS) induced by 50-min left middle cerebral artery occlusion. Adult-male Sprague-Dawley rats (n = 60) were divided equally into group 1 (sham-control), group 2 (AIS), group 3 [AIS-ADMSC (1.2×106 cells)], group 4 [AIS-exosome (100μg)], and group 5 (AIS-exosome-ADMSC). All therapies were provided intravenously at 3h after AIS procedure. BIZ determined by histopathology (by day-60) and brain MRI (by day-28) were highest in group 2, lowest in group 1, higher in groups 3 and 4 than in group 5, but they showed no difference between groups 3 and 4 (all p < 0.0001). By day-28, sensorimotor functional results exhibited an opposite pattern to BIZ among the five groups (p < 0.005). Protein expressions of inflammatory (inducible nitric oxide synthase/tumor necrosis factor-α/nuclear factor-κB/interleukin-1β/matrix metalloproteinase-9/plasminogen activator inhibitor-1/RANTES), oxidative-stress (NOX-1/NOX-2/oxidized protein), apoptotic (caspase-3/ Poly-ADP-ribose polymerase), and fibrotic (Smad3/transforming growth factor-β) biomarkers, and cellular expressions of brain-damaged (γ-H2AX+/ XRCC1-CD90+/p53BP1-CD90+), inflammatory (CD11+/CD68+/glial fibrillary acid protein+) and brain-edema (aquaporin-4+) markers showed a similar pattern of BIZ among the groups (all n < 0.0001). In conclusion, xenogenic ADMSC/ADMSC-derived exosome therapy was safe and offered the additional benefit of reducing BIZ and improving neurological function in rat AIS.

Cyclosporine-assisted adipose-derived mesenchymal stem cell therapy to mitigate acute kidney ischemia–reperfusion injury

IntroductionThis study tested the hypothesis that cyclosporine (CsA)-supported syngeneic adipose-derived mesenchymal stem cell (ADMSC) therapy offered superior attenuation of acute ischemia–reperfusion (IR) kidney injury to either therapy alone.MethodsAdult Sprague–Dawley rats (n = 40) were equally divided into group 1 (sham controls), group 2 (IR injury), group 3 (IR + CsA (20 mg/kg at 1 and 24 hours after procedure)), group 4 (syngeneic ADMSC (1.2×106) at 1, 6 and 24 hours after procedure), and group 5 (IR + CsA-ADMSC).ResultsBy 72 hours after the IR procedure, the creatinine level and the ratio of urine protein to creatinine were highest in group 2 and lowest in group 1, and significantly higher in groups 3 and 4 than in group 5 (all P <0.05 for inter-group comparisons), but showed no differences between groups 3 and 4 (P >0.05). The inflammatory biomarkers at mRNA (matrix metalloproteinase-9, RANTES, TNF-α), protein (TNF-α, NF-κB, intercellular adhesion molecule-1, platelet-derived growth factor), and cellular (CD68+) levels of IR kidney showed a similar pattern compared with that of creatinine in all groups (all P <0.05 for inter-group comparisons). The protein expressions of oxidative stress (oxidized protein), reactive oxygen species (NADPH oxidases NOX-1, NOX-2), apoptosis (Bcl-2–associated X protein, caspase-3 and poly(ADP-ribose) polymerase) and DNA damage (phosphorylated H2A histone family member X-positive, proliferating cell nuclear antigen-positive cells) markers exhibited a pattern similar to that of inflammatory mediators amongst all groups (all P <0.05 for inter-group comparisons). Expressions of antioxidant biomarkers at cellular (glutathione peroxidase, glutathione reductase, heme oxygenase-1 (HO-1)) and protein (NADPH dehydrogenase (quinone)-1, HO-1, endothelial nitric oxide synthase) levels, and endothelial progenitor cell markers (C-X-C chemokine receptor type 4-positive, stromal cell-derived factor-1α-positive) were lowest in groups 1 and 2, higher in groups 3 and 4, and highest in group 5 (all P <0.05 for inter-group comparisons).ConclusionCombination therapy using CsA plus ADMSCs offers improved protection against acute IR kidney injury.

Administered circulating microparticles derived from lung cancer patients markedly improved angiogenesis, blood flow and ischemic recovery in rat critical limb ischemia

BackgroundWe hypothesized that lung cancer patient’s circulating microparticles (Lc-MPs) could promote angiogenesis, blood flow in ischemic zone and ischemic recovery in rat critical limb ischemia (CLI).MethodsTo investigate the impact of MP therapy on reversing the setting of CLI, adult-male Sprague–Dawley rats (n=50) equally randomized into sham control (SC) (group 1), SC-Lc-MPs (1.0 x 107 particles) (group 2), CLI (group 3), CLI-Hs-MPs (MPs from healthy-subject) (group 4), and CLI-Lc-MPs (group 5) were sacrificed by post-CLI day-14.ResultsIn vitro study showed that Lc-MPs enhanced VEGFR2 expression, angiogenesis, nitric-oxide production, and endothelial cell proliferation (all p<0.005). By days 7 and 14, Laser Doppler showed significantly higher ischemic/normal blood-flow ratio in groups 1 and 2 compared with group 3, and was significantly higher in group 4 and further elevated in group 5 (p<0.0001). Numbers of small vessels and endothelial markers (CD31+ and vWF+ cells) and protein expressions (eNOS, CD31) exhibited a pattern identical to Lasre Doppler among the five groups (all p<0.001). Pro-angiogenic factors (VEGF, CXCR4, SDF-1α, HGF) at cellular and protein levels showed a significant step-wise increase from groups 1 and 2 to groups 3, 4, and 5 (all p<0.001). Protein expressions of fibrotic (Smad3, TGF-β) and apoptotic (mitochondrial Bax, cleaved caspase 3, and PARP) biomarkers displayed an opposite pattern compared to that of Laser Doppler, whereas the protein expressions of anti-fibrotic (Smad1/5, BMP-2) and anti-apoptotic (Bcl-2) biomarkers showed an identical pattern compared with that of Laser Doppler among groups 1 to 3, and 5 (all p<0.001).ConclusionAdministration of Lc-MPs augmented angiogenesis and restored blood flow in a rat of CLI.

Levels of Circulating Microparticles in Patients with Chronic Cardiorenal Disease

AIM Cardiac and renal diseases are common disorders that frequently coexist. We tested the hypothesis that the levels of circulating endothelial-derived apoptotic microparticles (EDA-MPs; CD31(＋)CD42b(-)AN(-)V(＋)) and platelet-derived apoptotic microparticles (PDA-MPs; CD31(＋)CD42b(＋)AN(-)V(＋)) are useful biomarkers for predicting the presence of cardiorenal disease (CRD). METHODS A total of 68 patients with chronic kidney disease (CKD) and angina pectoris (CKD-AP) undergoing cardiac catheterization were prospectively enrolled into group 1, 10 patients with coronary artery disease (CAD) without CKD were enrolled into group 2 (CAD(＋)CKD(-)) and 10 patients without CAD and CKD were enrolled into group 3 (CAD(-)CKD(-)). RESULTS The serum creatinine levels were significantly higher, whereas the estimated glomerular filtration rates (eGFRs) were significantly lower, in group 1 than in the other two groups (all p < 0.02). The circulating levels of EDA-MPs and PDA-MPs did not differ between the patients with and without CKD (all p > 0.2). However, the circulating levels of EDA-MPs and PDA-MPs were significantly higher in group 2 than in groups 1 and 3 (all p < 0.03). In addition, differences were noted in the circulating EDA-MP and PDA-MP levels between groups 1 and 3, although without statistical significance (all p > 0.09). Meanwhile, among the CKD patients, the subgroup analysis showed that the levels of MPs were significantly higher in those with CAD than in those without (all p=0.001), while a multivariate analysis demonstrated that CAD was the only factor independently predictive of high levels of circulating EDA-MPs and PDA-MPs (p=0.033). CONCLUSIONS The link with increased circulating levels of MPs is more consistent in patients with CAD than in those with CKD.

Intra-carotid arterial administration of autologous peripheral blood-derived endothelial progenitor cells improves acute ischemic stroke neurological outcomes in rats.

OBJECTIVE We tested the hypothesis that transfusion of autologous peripheral blood-derived endothelial progenitor cells (PBDEPC) via the internal carotid artery could reduce brain-infarct zone (BIZ) and neurological deficit in rats following acute ischemic stroke (IS) induced by 50-min left middle cerebral artery occlusion. DESIGN Adult male Sprague-Dawley rats (n=60) were equally divided into group 1 [sham control (SC)], group 2 [SC-PBDEPC (5.7 × 10(6)/kg)], group 3 (IS), group 4 [IS-low-dose PBDEPC (1.7 × 10(6)/kg)], group 5 [IS-high-dose PBDEPC (5.7×10(6)/kg)]. Groups 2 to 5 received G-CSF (35 μg/kg subcutaneously) for 4 days before drawing blood for PBDEPC culture. MEASUREMENTS AND MAIN RESULTS By day 90, BIZ determined by histopathology (area) and brain MRI (volume) were highest in group 3, lowest in groups 1 and 2, higher in group 4 than in group 5 (all p<0.0001), and not significantly different between groups 1 and 2. Sensorimotor functional results exhibited an opposite pattern of BIZ among groups 3 to 5 (p<0.005). Angiogenesis biomarkers (SDF-1α, CXCR4, VEGF, angiopoietin-1) significantly increased progressively from groups 1 and 2 to group 5 (all p<0.0001). Oxidative-stress (NOX-1, NOX-2, oxidized protein), apoptotic (cleaved caspase 3 and PARP, mitochondrial Bax), inflammatory (MMP-9, TNF-α, AQP-4, GFAP, iNOS), and brain-damaged (cytosolic cytochrome-C) biomarkers showed an identical pattern, whereas anti-inflammatory (Bcl-2), mitochondrial preservation (mitochondrial cytochrome-C, PGC-1α), and endothelial function (CD31+, vWF+, eNOS) biomarkers, and vessel density showed an opposite pattern of BIZ among these five groups (all p<0.001). CONCLUSION Higher-dose was superior to lower-dose EPC treatment for reducing BIZ and improving neurological functional outcome.

Role of multiple free flaps in head and neck reconstruction.

Purpose of reviewThe benefit of using multiple simultaneous free flaps for postablative extensive composite head and neck reconstruction has gradually become increasingly accepted worldwide, with recent case series being reported from several continents. This review summarizes the cogent conclusions that can be drawn from this growing international experience. Recent findingsRunning themes include: firstly, that careful organization of the operation is critical to expediency to the extent that double and single free flap reconstructions are not greatly different in duration; secondly, that the functional results of double free flap reconstructions are generally better than single free flap reconstructions when the defect is appropriately extensive; and thirdly, that there remains a lack of consensus regarding which patients would and would not benefit from a double free flap approach from a survival perspective. SummaryLocal audit should guide local practice for when double free flap reconstructions are or are not appropriate, as survival data are greatly variable internationally because of the differences in treatment, margins status for resections and, amongst other factors, cause and aggression of tumors. This is especially the case for the most infiltrative malignancies that mandate extensive composite resections for which a double free flap procedure would likely provide the best long-term functional and aesthetic results.