Lower Limb Perfusion Asymmetries in Humans at Rest and Following Activity—A Collective View

The significance of lower limb perfusion asymmetries remains unknown in healthy individuals. Our study aims to understand how factors such as posture, sex, age, and body weight relate to perfusion. Data from studies previously published by our group, including experiments using laser Doppler flowmetry as the gold standard for perfusion measurements in baseline, (various) challenge, and recovery phases was assembled from a total of 139 healthy participants. Body position was shown to be a primary determinant of perfusion asymmetry, especially in women. Effects of sex and age were also analyzed. In a supine position, perfusion asymmetries seemed to relate negatively in the aged group of participants, where challenge and recovery seemed to follow different processes. In the upright position, young men and women have shown comparable distributions and asymmetry ratios at baseline and recovery. In the aged group, differences between sexes were observed at baseline, but again, the course of the asymmetry ratios with challenge was essentially similar in men and women. Our analysis suggests that ageing is a critical determinant in our upright study sample, as higher baseline asymmetries and longer recoveries after challenge were linked in older males with higher body mass index (BMI).

The diagnostic value of modified systemic ınflammation score in predicting post-operative outcomes of cutaneous melanoma patients who underwent ısolated limb perfusion

Background In-transit metastasis is considered a locoregional disease in cutaneous melanoma (CM) patients. Isolated limb perfusion (ILP) is among the treatment options in selected cases. The aim of this study was to determine the success of pre- and post-perfusion mSIS values in predicting the potential complications and the prognosis of the disease by investigating the early and long-term results of mSIS values calculated before and after ILP in CM cases with in-transit metastases. Materials and methods Patients who underwent ILP within the period from 2014 to 2020 in our department were retrospectively scanned. A total of 20 patients were found to undergo ILP. The scores obtained from modified inflammation score (mSIS) were formulated according to albumin (Alb) and lymphocyte to monocyte ratio (LMR) scores. Results The mean follow-up time was 20.47 months. Complications requiring surgical intervention developed in three patients. According to the Wieberdink local toxicity classification, the majority (70%) of the patients were found to be grade II. Based on pre-perfusion mSIS values, 8 patients were classified as mSIS 0 while six patients were classified as mSIS 1 and 2. Based on post-perfusion mSIS values, 14 patients and one patient were classified as mSIS 2 (70%) and mSIS 0, respectively. Accordingly, univariate analysis showed that mSIS 1 and mSIS 2 were negative prognostic factors for mean survival in the pre-perfusion period (HR 0.162, 95% CI 0.036–0.729; p = 0.018 and HR: 0.223, 95% CI 0.049–1.019; p = 0.053) whereas albumin (Alb) and lymphocyte to monocyte ratio (LMR) were not independent prognostic factors for mean survival. Conclusion The mSIS values calculated in the pre-perfusion period can give an opinion about the OS of the patients whereas post-perfusion mSIS values may predict potential surgical complications and local toxicities.

Indocyanine Green for Leakage Control in Isolated Limb Perfusion

Sarcomas are characterized by a high metastatic potential and aggressive growth. Despite surgery, chemotherapy plays an important role in the treatment of these tumors. Optimal anti-cancer therapy with maximized local efficacy and minimized systemic side effects has been the object of many studies for a long time. To improve the local efficacy of anti-tumor therapy, isolated limb perfusion with high-dose cytostatic agents has been introduced in surgical oncology. In order to control the local distribution of substances, radiolabeled cytostatic drugs or perfusion solutions have been applied but often require the presence of specialized personnel and result in a certain exposure to radiation. In this study, we present a novel strategy using indocyanine green to track tumor perfusion with high-dose cytostatic therapy. In a rat cadaver model, the femoral vessels were cannulated and connected to a peristaltic pump to provide circulation within the selected limb. The perfusion solution contained indocyanine green and high-dose doxorubicin. An infrared camera enabled the visualization of indocyanine green during limb perfusion, and subsequent leakage control was successfully performed. Histologic analysis of sections derived proximally from the injection site excluded systemic drug dispersion. In this study, the application of indocyanine green was proven to be a safe and cost- and time-efficient method for precise leakage control in isolated limb perfusion with a high-dose cytostatic agent.

Role of Isolated Limb Perfusion in the Era of Targeted Therapies and Immunotherapy in Melanoma. A Systematic Review of The Literature

Background. Isolated limb perfusion (ILP) is a locoregional procedure indicated by the unresectable melanoma of the limbs. Its complexity and highly demanding multidisciplinary approach means that it is a technique only implemented in a few referral centers around the globe. This report aims to examine its potential role in the era of targeted therapies and immunotherapy by conducting a systematic review of the literature on ILP. Methods. PubMed, Embase and Cochrane Library were searched. The eligibility criteria included publications from 2000–2020 providing valid data o effectiveness, survival or toxicity. Studies in which the perfusion methodology was not clearly described, letters to the editor, non-systematic reviews and studies that applied outdated clinical guidelines were excluded. To rule out studies of a low methodological quality and assess the risk of bias, the following aspects were also required: a detailed description of the applied ILP regimen, the clinical context, follow-up periods, analyzed clinical endpoints, and the number of analyzed ILPs. The disagreements were resolved by consensus. The results are presented in tables and figures. Results. Twenty-seven studies including 2637 ILPs were selected. The median overall response rate was 85%, with a median complete response rate of 58.5%. The median overall survival was 38 months, with a 5-year overall survival of 35%. The toxicity was generally mild according to Wieberdink toxicity criteria. Discussion. ILP still offer a high efficacy in selected patients. The main limitation of our review is the heterogeneity and age of most of the articles, as well as the absence of clinical trials comparing ILP with other procedures, making it difficult to transfer its results to the current era. Conclusions. ILP is still an effective and safe procedure for selected patients with unresectable melanoma of the limbs. In the era of targeted therapies and immunotherapy, ILP remains an acceptable and reasonable palliative treatment alternative, especially to avoid limb amputations. The ongoing clinical trials combining systemic therapies and ILP will provide more valuable information in the future to clarify the potential synergism of both strategies.

Chemical and Functional Separation of Pro- and Antiangiogenic Components of Danhong Injection in Ischemic or Tumor Vascular Models

Objective: We aimed to investigate the chemical basis and mechanism of angiogenesis regulation by a multicomponent Chinese medicine Danhong injection (DHI). Methods: A chemical fraction library of DHI was screened and validated for angiogenesis activities by tube formation and migration assays. Mouse ischemic and tumor vascular models were used to verify the angiogenesis effects in vivo. Migration ability of the main monomers of proangiogenic component (PAC) and antiangiogenic component (AAC) in EA.hy926 cells were determined by migration assay. qPCR analyses were performed to access whether the main monomers of PAC or AAC could affect the expression of angiogenesis-related genes in ECs. Western blotting was used to verify the main monomers PAC and AAC effects on CXCR4 protein expression. Results: Two chemically-distinct fractions, including promotion and inhibition of angiogenesis, were identified in DHI. PAC enhanced angiogenesis and improved recovery of ischemic limb perfusion while AAC reduced Lewis lung tumor growth in vivo in VEGFR-2-Luc mice. CA and RA upregulated the expression of TSP1 and downregulated the expression of KDR and PECAM genes. CXCR4 expression was significantly decreased by CA and RA, but increased by PAI, consistent with their differential effects on EC migration. Conclusion: DHI is capable of bi-directional regulation of angiogenesis in a disease-specific manner. The proangiogenesis activity of DHI promotes ischemic vascular injury repair, whereas the anti-angiogenesis activity inhibits tumor growth. The best pro- and anti-angiogenesis activities are composed of unique chemical combinations that differentially regulate angiogenesis-related gene network.

Evaluation of totally implantable catheters in healthy horses

Background For horses requiring prolonged daily cephalic intravenous regional limb perfusion (IVRLP), the use of a totally implantable catheter (TIC) could be indicated to reduce complications associated with frequent venipuncture or external catheterization. This study aims to evaluate the implantation technique of the TIC in the cephalic vein of horses for IVRLP, describe the complications associated with the device’s placement and use, and assess its viability up to 60 days after implantation. Totally implantable catheters, cut to 15 cm (n = 5) and 46 cm (n = 5) in length, were implanted into one cephalic vein in ten adult horses (n = 10). Twenty-four hours following placement, IVRLP with contrast was performed via the TIC and evaluated with radiography. Physical examinations, lameness evaluation, hematologic assessment, and the catheter patency tests were performed at scheduled intervals for the duration of catheterization (7–60 days). Results Catheters were implanted without difficulty and allowed for IVRLP 24 h post implantation. Complications resulted in removal of the catheters, with four maintained for 7 days, three in place for 15 days, and three catheters maintained for 60 days. Complications included lameness, limb swelling, catheter kinking, and venous thrombosis. Conclusions The implantation technique of the TIC in the cephalic vein of horses is feasible and requires minimal technical effort. Although TIC allows venous access without the need for repeated venipuncture, its long-term use presents complications. For horses requiring prolonged daily cephalic IVRLP, the use of a TIC could be indicated. However, the high incidence of venous thrombosis may limit clinical application.

A Thermoresponsive Chitosan/β-Glycerophosphate Hydrogel for Minimally Invasive Treatment of Critical Limb Ischaemia

A reduction in blood supply to any limb causes ischaemia, pain and morbidity. Critical limb ischaemia is the most serious presentation of peripheral vascular disease. One in five patients with critical limb ischaemia will die within six months of diagnosis and one in three will require amputation in this time. Improving blood flow to the limb, via the administration of angiogenic agents, could relieve pain and avoid amputation. Herein, chitosan is combined with β-glycerophosphate to form a thermoresponsive formulation (chitosan/β-GP) that will flow through a syringe and needle at room temperature but will form a gel at body temperature. The chitosan/β-GP hydrogel, with or without the angiogenic molecule desferrioxamine (DFO), was injected into the mouse hind limb, following vessel ligation, to test the ability of the formulations to induce angiogenesis. The effects of the formulations were measured using laser Doppler imaging to determine limb perfusion and CD31 staining to quantify the number of blood vessels. Twenty-eight days following induction of ischaemia, the chitosan/β-GP and chitosan/β-GP + 100 µM DFO formulations had significantly (p < 0.001 and p < 0.05, respectively) improved blood flow in the ischaemic limb compared with an untreated control. Chitosan/β-GP increased vessel number by 1.7-fold in the thigh of the ischaemic limb compared with an untreated control, while chitosan/β-GP + 100 µM DFO increased vessel number 1.8-fold. Chitosan/β-GP represents a potential minimally invasive treatment for critical limb ischaemia.

MicroRNA-126 mimic administration accelerates vascular perfusion recovery and angiogenesis in a hind limb ischemia model

Background Peripheral arterial disease caused mainly by atherosclerosis portent significant morbidity, adverse prognosis and mortality, with localized treatment approaches aiming at symptom alleviation and improvement of circulation. Recently, scientific interest has been shifted towards epigenomics, with microRNAs appearing as a future therapeutic target in ischemic cardiovascular diseases due to their potential in regulating angiogenesis. Purpose We investigated the pro-angiogenic effect of miRNA-126 mimic in an in vivo model of hind limb ischemia. Methods Ten-week-old male C57Bl/6 mice (n=20) were subjected to left femoral artery ligation and were treated with microRNA-126 mimic at a dose of 5mg/kg (Group A, n=10) or 0.2ml normal saline (Group B, n=10) on days 1, 3 and 7. Laser Doppler imaging was performed to verify successful ligation on day 0 and to evaluate differences in the ischemic-to-normal (I/N) hind limb perfusion ratio on day 7 and 28. Muscle tissue expression of microRNA-126 and vascular endothelial growth factor (VEGF) was determined via PCR. Results Following microRNA-126 mimic administration in Group A subjects, we noted a qualitative and quantitative stepwise increase in I/N hind limb perfusion ratio [Day 0: 0.354 (0.276, 0.455) vs. Day 8: 0.775 (0.700, 0.844) vs. Day 28: 0.681 (0.660, 0.896), p=0.001] (Figure 1, Panels A and B). In Group B a stepwise increase of lesser magnitude was observed in I/N hind limb perfusion ratio [Day 0: 0.267 (0.164, 0.383) vs. Day 8: 0.400 (0.338, 0.418) vs. Day 28: 0.539 (0.483, 0.603), p=0.074]. Importantly, over time changes of I/N hind limb perfusion ratio were significantly higher in group A compared to group B (p for interaction=0.005) (Figure 1, Panel B). Muscle tissue expression of microRNA-126 in the ischemic hind limb of Group A was 350-fold lower compared to the ischemic hind limb of Group B (p&lt;0.001) (Figure 1, Panel C). A higher expression (14.2-fold) of VEGF in the ischemic hind limb of microRNA-126-treated mice compared to that of control group was detected (p&lt;0.001) (Figure 1, Panel C). A statistically significant negative correlation was noted between microRNA-126 and VEGF tissue expression levels in the ischemic limbs of both Group A and B subjects whereas no correlation between microRNA-126 and VEGF was observed in the non-ischemic hind limbs of the entire study population (Figure 1, Panel D). Conclusion MicroRNA-126 mimic delivery in the ischemic hind limb of mice can accelerate vascular perfusion recovery via angiogenesis, which is mediated by VEGF expression. FUNDunding Acknowledgement Type of funding sources: None. Figure 1