Hydroxyurea and fetal hemoglobin effect on leg ulcers in patients with sickle cell disease.

The presence of leg ulcers in individuals with sickle cell disease often represents an early sign of vasculopathy and future end organ damage. Pathophysiological mechanisms of formation and evolution of leg ulcers are poorly understood; nevertheless, HbF has been associated with lower incidence of leg ulcers, while hydroxyurea has been correlated with high risk of leg ulcers. As a result, there is hesitation regarding hydroxyurea use in patients with SCD and leg ulcers. In this study, we aim to define (1) a target of HbF that offers protection against leg ulcer development and (2) the impact of hydroxyurea therapy on leg ulcer prevalence. Our study demonstrated that in order to reduce leg ulcer incidence by one-third, a HbF > 25% is needed, a threshold not commonly reached and maintained in the adult SCD population. Importantly, leg ulcer incidence appears to be independent of HU use (p = 0.50). Our interpretation of this data is that the use of HU in a patient with SCD and leg ulcers should be guided by a careful assessment of risks and benefits of this therapeutic modality.

Effect of Vitamin D3 in combination with Omega-3 Polyunsaturated Fatty Acids on NETosis in Type 2 Diabetes Mellitus Patients.

An understanding of the consequences of oxidative/halogenative stress triggered by neutrophil activation is impossible without considering NETosis. NETosis, formation of neutrophil extracellular traps (NETs), is known to promote microthrombus formation and impair wound healing in type 2 diabetes mellitus (T2DM) patients. Therefore, there is a need to search for drugs and treatment approaches that could prevent excessive NET formation. We aimed to evaluate the effect of vitamin D3 in combination with omega-3 polyunsaturated fatty acids (vitamin D3/omega-3 PUFAs) on NETosis in T2DM patients with purulent necrotizing lesions of the lower extremities. Patients and healthy subjects had vitamin D3 deficiency. Patients received, beyond standard treatment, 6000 IU of vitamin D3 and 480 mg of omega-3 PUFAs, and healthy subjects 1000 IU of vitamin D3 and 240 mg of omega-3 PUFAs daily for seven days. Neutrophil activation in ex vivo blood by phorbol-12-myristate-13-acetate (PMA) was used as a NETosis model. The percentage of blood NETs relative to leukocytes (NETbackground) before vitamin D3/omega-3 PUFA supplementation was 3.2%-4.9% in healthy subjects and 1.7%-10.8% in patients. These values rose, respectively, to 7.7%-9.1% and 4.0%-17.9% upon PMA-induced NETosis. In addition, the leukocyte count decreased by 700-1300 per 1 µL in healthy subjects and 700-4000 per 1 µL in patients. For both patients and healthy subjects, taking vitamin D3/omega-3 PUFAs had no effect on NETbackground but completely inhibited PMA-induced NET formation, though neutrophils exhibited morphological features of activation. Also, leukocyte loss was reduced (to 500 per 1 µL). For patients on standard treatment alone, changes occurred neither in background NETs and leukocytes nor in their amount after PMA stimulation. The decreased ability of neutrophils to generate NETs, which can be achieved by vitamin D3/omega-3 PUFA supplementation, could have a positive effect on wound healing in T2DM patients and reduce the incidence and severity of complications.

Correlation between vitamin D levels and hard-to-heal wounds: a systematic review.

OBJECTIVE: Hard-to-heal wounds are a major biological, psychological, social and financial burden on both individual patients and the broader health system. They are associated with a variety of comorbidities and have a complex aetiology, but are typically associated with nutritional deficiencies, such as low vitamin D levels. This systematic literature review evaluates the current research regarding the connection between inadequate vitamin D status and wound healing. METHOD: PubMed and EBSCO databases were searched following PRISMA guidelines for primary research studies relating to pressure ulcers, diabetic ulcers or venous ulcers and vitamin D status. RESULTS: A total of 10 studies, involving 2359 participants, met the inclusion criteria. There was a strong correlation between low levels of 25-hydroxyvitamin D and the presence of all three types of hard-to-heal wounds. CONCLUSION: Research suggests a correlation between low vitamin D levels and hard-to-heal wounds. However, it is not clear if the relationship is causal or only correlational. There is also emerging evidence on the use of vitamin D supplementation for the treatment of hard-to-heal wounds. More research is needed to understand the correlation between vitamin D and hard-to-heal wounds.

Leukemoid reaction causing arterial thrombus in a patient with lung adenocarcinoma.

A leukemoid reaction is typically defined as white blood cell (WBC) count >50×109/L, predominantly neutrophil precursors, that are not due to tumour involvement in the bone marrow and not derived from clones. Leukemoid reactions associated with malignancy, known as paraneoplastic leukemoid reactions, are less common and are most notably seen with non-small cell lung cancer. A 64-year-old woman presented with right leg painful ulceration. On examination, she had multiple venous stasis ulcers more severe on the right, with no palpable pulses in her lower extremities. Her WBC count was 124×109/L and platelets were 517×109/L. Arterial dopplers showed limb-threatening arterial insufficiency which prompted right femoral endarterectomy. Few months earlier she was diagnosed with metastatic lung adenocarcinoma to the bone and she had leukemoid reaction with WBC 43.920× 109/L with 90% neutrophils. Repeat imaging showed progression of her malignancy and she passed shortly after. Inflammation is a key element of carcinogenesis and cancer progression. Among the different tumours, lung cancer is a non-haematologic malignancy that is most closely associated with leucocytosis. Some studies have found that leucocytosis was significantly associated with metastasis and shorter survival irrespective of other factors such as age or sex. The mechanism remains unclear however elevated levels of granulocyte colony-stimulating factor (CSF), granulocyte macrophage-CSF and interleukin 6 have been linked to this phenomena. The degree of leucocytosis seen in our patient is suggestive of CSF production leading to a paraneoplastic leukemoid reaction.

A study of the geographic distribution and associated risk factors of leg ulcers within an international cohort of sickle cell disease patients: the CASiRe group analysis.

Vasculopathy is a hallmark of sickle cell disease ultimately resulting in chronic end organ damage. Leg ulcer is one of its sequelae, occurring in ~ 5-10% of adult sickle cell patients. The majority of leg ulcer publications to date have emanated from single center cohort studies. As such, there are limited studies on the geographic distribution of leg ulcers and associated risk factors worldwide. The Consortium for the Advancement of Sickle Cell Research (CASiRe) was formed to improve the understanding of the different phenotypes of sickle cell disease patients living in different geographic locations around the world (USA, UK, Italy, Ghana). This cross-sectional cohort sub-study of 659 sickle cell patients aimed to determine the geographic distribution and risk factors associated with leg ulcers. The prevalence of leg ulcers was 10.3% and was associated with older age, SS genotype, male gender, and Ghanaian origin. In fact, the highest prevalence (18.6%) was observed in Ghana. Albuminuria, proteinuria, increased markers of hemolysis (lower hemoglobin, higher total bilirubin), lower oxygen saturation, and lower body mass index were also associated with leg ulceration. Overall, our study identified a predominance of leg ulcers within male hemoglobin SS patients living in sub-Saharan Africa with renal dysfunction and increased hemolysis.

High levels of proinflammatory cytokines IL-6 and IL-8 are associated with a poor clinical outcome in sickle cell anemia.

Sickle cell anemia (SCA) pathophysiology is characterized by the activation of sickle red blood cells, reticulocytes, leukocytes, platelets, and endothelial cells, and with the expression of several inflammatory molecules. Therefore, it is conceivable that variations in levels of proinflammatory cytokines may act as a signaling of differential clinical course in SCA. Here, we evaluated the clinical impact of proinflammatory cytokines interleukin 1-ß (IL-1ß), interleukin 6 (IL-6), and interleukin 8 (IL-8) in 79 patients with SCA, followed in a single reference center from northeastern Brazil. The main clinical/laboratory data were obtained from patient interview and medical records. The proinflammatory markers IL-1ß, IL-6, and IL-8 were evaluated by using commercially available enzyme-linked immunosorbent assay kits. According to levels of the proinflammatory markers, we observed that patients who had a higher frequency of VOC per year (P = 0.0236), acute chest syndrome (P = 0.01), leg ulcers (P = 0.0001), osteonecrosis (P = 0.0006), stroke (P = 0.0486), and priapism (P = 0.0347) had higher IL-6 levels compared with patients without these clinical complications. Furthermore, increased levels of IL-8 were found in patients who presented leg ulcers (P = 0.0184). No significant difference was found for IL-1ß levels (P > 0.05). In summary, the present study emphasizes the role of inflammation in SCA pathophysiology, reveals an association of IL-8 levels and leg ulcer occurrence, and indicates that IL-6 levels can be used as a useful predictor for poor outcomes in SCA.

Characterization of the local wound environment following treatment of chronic leg ulcers with SIS wound matrix.

BACKGROUND: The SIS Wound Matrix (SISWM) has been shown to improve healing of chronic ulcers over standard of care. In this study, we tested the hypothesis that chronic venous ulcers responsive to treatment with SISWM would more closely mimic an acute wound state as opposed to unresponsive ulcers. METHODS: Serum and wound exudate were collected at baseline and then weekly for up to 12 weeks from 12 patients receiving multiple applications of the SISWM. Levels of matrix metalloproteinases (MMP-1, MMP-2, MMP-3, MMP-9, and MMP-12), pro-inflammatory cytokines (IL-1ß, TNF-α, IL-8), and transforming growth factor beta (TGF-ß1) were evaluated. A variety of Th1/Th2 cytokines were also assayed, as were systemic anti-SIS and anti-α-gal antibody titers. RESULTS: Seven of the 12 patients eventually healed their wounds. Results showed significant decreases in MMP-1, MMP-2, MMP-3, MMP-9, TNF-α and IL-8, and significant increases in TGF-ß1 in wounds responding to treatment with the SISWM versus wounds that did not respond to treatment. None of the 12 patients formed a measurable serum antibody response to the SISWM. CONCLUSIONS: These data show that SISWM does not lead to immune system recognition or sensitization to the matrix and that wounds that went on to heal following treatment were characterized by a more acute wound state. The study confirms that the wound environment is important to healing and that turning a wound toward an acute biochemical state is key to the healing process.

Multifactorial Painful Leg Ulcers Due to Hyperhomocysteinemia, Plasminogen Activator Inhibitor-1 4G/5G Heterozygote Gene Mutation, and Beta Thalassemia Minor: A Case Report.

Leg ulcers may occur due to many autoimmune, hereditary, inflammatory, and infectious causes including venous, arterial, and neuropathic ulcers. Hyperhomocysteinemia is a metabolic disorder caused by various enzyme defects in methionine metabolism. The most common cause is methylenetetrahydrofolatreductase (MTHFR) enzyme gene mutations. Hyperhomocysteinemia is an independent risk factor for deep vein thrombosis and peripheral arterial disease. The effects of endothelial cell damage on smooth muscle hypertrophy, platelet aggregation, coagulation, and fibrinolysis cause atherogenesis and thrombosis, leading to venous and arterial lower extremity ulcers. In this article, we report the case of a 47-year-old male patient who was admitted to our clinic due to painful leg ulcers that started 1 year ago. He had a history of vena cava inferior thrombosis, deep vein thrombosis, and 40 pack-year smoking. Histopathological examination of punch biopsy taken from ulcerative lesion showed intense inflammatory infiltration in the middle dermis, erythrocyte extravasation, leukocytoclasia, and thrombus formation in a small diameter venule lumen. There were nonspecific findings in direct immunofluorescence examination. He was found as having MTHFR C677T homozygote and plasminogen activator inhibitor-1 4G/5G heterozygote gene mutation with high homocysteine level of 22.90 µmol/L, and he was diagnosed as hyperhomocysteinemia. He was recommended to quit smoking because it triggered thrombosis in hyperhomocysteinemia. Herein, we present a case of hyperhomocysteinemia due to MTHFR mutation, which is one of the rare hereditary thrombophilia causes.

Low nitric oxide level is implicated in sickle cell disease and its complications in Ghana.

BACKGROUND: Nitric oxide (NO) plays a fundamental role in maintaining normal vasomotor tone. Recent clinical and experimental data suggest that NO may play a role in the pathogenesis and therapy of sickle cell disease (SCD). The aim of this study was to determine NO metabolites (NOx) in SCD patients at steady state and in vaso-occlusive crisis (VOC), as well as those with hemolytic clinical sub-phenotype that includes leg ulcers and priapism. METHODOLOGY: This was a case-control cross-sectional study conducted on a total of 694 subjects including 148 comparison group HbAA, 208 HbSS SCD patients in steady state, 82 HbSC SCD patients in steady state, 156 HbSS SCD patients in VOC, 34 HbSC SCD patients in VOC, 34 HbSS SCD patients in post VOC, 21 HbSS SCD patients with leg ulcer and 11 HbSS SCD patients with priapism, with age ranging from 15 to 65 years. Laboratory diagnosis of SCD was done at the Sickle Cell Clinic of the Korle-Bu Teaching Hospital. Plasma nitric oxide metabolites were measured using Griess reagent system by ELISA method. RESULTS: Mean NOx of 59.66±0.75 µMol/L in the comparison group was significantly different from those in steady state (P=0.02). During VOC, there was a significant reduction in mean NOx levels to 6.08±0.81 µMol/L (P<0.001). Mean NOx levels were however, significantly higher (50.97±1.68 µMol/L) (P<0.001) in the immediate postcrisis period. The mean NOx levels in the leg ulcer (21.70±1.18 µMol/L) (P<0.001) and priapism (28.97±1.27 µMol/L) (P<0.001) patients were significantly low as compared to the SCD patients in the steady state and comparison group. CONCLUSION: This study presents the first report on plasma NOx levels in SCD complication in Ghanaian SCD patients and confirms reduced plasma NOx levels in SCD patients in general.

Evaluation of sociodemographic, clinical, and laboratory markers of sickle leg ulcers among young nigerians at a tertiary health institution.

BACKGROUND: Sickle leg ulcer (SLU) is a chronic and debilitating complication of sickle cell disease (SCD) associated with huge physical and psychosocial discomfort. The occurrence of SLU has remained steady despite successful preventive strategies and advances in SCD care. Although multifactorial factors have been implicated in SLU, these are not fully understood, and data on how these relate to young Nigerian SCD patients are scanty. AIMS: This study aims to evaluate the sociodemographic, clinical, and laboratory markers of SLU in a young Nigerian SCD cohort. PATIENTS AND METHODS: This study involved 109 young SCD patients and 67 healthy peers. The sociodemographic and laboratory parameters of the participants were examined in addition to the evaluation of the SCD cohort for SLU. RESULTS: Only the HbSS patients had SLU. This was found in six of them giving a prevalence of 5.9% (6/101). Their median age was 17, range 14-21 years. There was a preceding history of trauma in 4 (66.7%), and this included a case of traditional scarifications for local therapeutic purposes. Two of the three (66.7%) males with SLU also had priapism (P = 0.0132). Patients with SLU were older, had less frequent bone pain crises, and significantly belonged to the low socioeconomic class (P < 0.05). Although patients with SLU had relatively higher lactate dehydrogenase, platelet count, aspartate transaminase, bilirubin, white blood cell, and lower Hb concentration and HbF, these did not attain statistical significance (P > 0.05). CONCLUSION: This study confirms that SLU is common among young SCD patients with HbSS genotype, low socioeconomic background, and older age. It also suggests that SLU could be more related to hemolysis-associated SCD phenotypes among the patients.

Autologous platelet rich plasma in the management of non-healing vasculitic ulcers.

AIM: To determine the role and efficacy of platelet rich plasma (PRP) in the healing of recalcitrant vasculitic ulcers and to study the rapidity of ulcer healing with respect to platelet concentration in the prepared PRP. METHODS: This is a pilot study which included 20 patients with biopsy-proven vasculitic ulcers after excluding human immunodeficiency virus, uncontrolled diabetes mellitus, hypertension, obesity, active smoking and platelet abnormalities in peripheral smear. PRP was prepared from whole blood by centrifugation and activated using 10% calcium chloride. Activated PRP was applied over the ulcer by spraying using a syringe. Weekly dressing was done until the ulcer healed and healing was monitored using the Leg Ulcer Measurement Tool (LUMT). RESULTS: The total number of patients were 20 (male four; female 16). The mean age was 34.5 ± 15.36 years (2.0 SD). Mean duration of illness was 8.6 ± 2.9 months(2.0 SD). Ulcers were located in the lower third of the leg (n = 13), ankle (n = 4) and foot (n = 3) in the descending order of frequency. On average, epithelization of the wound occurred at 4.4 weeks. Faster healing was seen in mixed connective tissue disease (average 3.5 weeks) and delayed healing in systemic lupus erythematosus (average 4.7 weeks). Average platelet count obtained in the PRP was 5.52 lakhs/mm3 to 18.6 lakhs/mm3 with an increase in platelet count of 3.4- to 6-fold, respectively. Duration of healing corresponded with the platelet concentration in the prepared PRP. All the 20 patients showed complete healing within 6 weeks. CONCLUSION: PRP is a safe, effective, outpatient procedure for the treatment of chronic recalcitrant vasculitic ulcers which improves the quality of life of patients and reduces their financial burden.

Red blood cell deformability is reduced in homozygous sickle cell disease patients with leg ulcers.

Previous reports differ as to whether a decreased elongation index (EI), a proxy for red blood cell (RBC) deformability, is associated with leg ulcers (LU) in people with homozygous sickle cell disease (SCD). We sought to determine whether erythrocyte deformability (ED) and haematological indices were associated with the presence of LU in patients with SCD. The study design was cross-sectional. Twenty-seven patients with LU and 23 with no history of ulceration were recruited into the study. A laser assisted rotational red cell analyzer was used in the determination of the EI. Haematological indices were determined using a CELL-DYN Ruby haematology analyzer. Data were normally distributed and presented as means±SD. Two-sample t-test was used to test for associations between haemorheological variables in SCD patients with and without LU. Statistical significance was taken as p < 0.05. The EI was significantly lower in the group with ulcers (0.30±0.07 vs. 0.35±0.07, p = 0.02). Haematological indices were comparable in patients with and without LU. Erythrocyte deformability, but not haematological indices, was associated with LU in patients with SCD.

Factors Affecting Mortality After Major Nontraumatic Lower Extremity Amputation.

Our aim was to evaluate the factors affecting the mortality of patients who underwent nontraumatic major lower limb amputation due to ischemic and/or diabetic causes. A total of 100 patients were included in the study. Among these patients, 70 (70%) underwent below-knee amputation, whereas 30 (30%) underwent above-knee amputation. Eleven (15.7%) of the 70 patients who underwent below-knee amputation and 12 (40%) of the 30 patients who underwent above-knee amputation (P = .008) were deceased. After multivariable Poisson regression analysis, female gender (risk ratio [RR] = 2.00, 95% CI = 1.07-3.74) and a neutrophil lymphocyte ratio (NLR) less than 6.8 (RR = 5.12, 95% CI = 1.86-14.08) were found to be independent risk factors for mortality. The value of 6.8 was used as a cutoff point for the NLR (area under the curve = 0.73, 95% CI = 0.62-0.85), with a sensitivity, specificity, positive predictive value, and negative predictive value of 83%, 66%, 57%, and 92%, respectively. The NLR and female gender were found to be independent factors that are related to increased mortality in patients who underwent lower limb amputation due to diabetic and/or ischemic causes. The coexistence of congestive heart failure and the amputation level (above knee) were found to be predictors of mortality in univariable analysis, but significance could not be demonstrated in multivariable analysis.

Critical Reviews: How we treat sickle cell patients with leg ulcers.

The past five decades have seen an improvement in the mortality and morbidity of sickle cell disease (SCD) because of prophylaxis against infectious complications, improved and expanded red cell transfusions, implementation of hydroxyurea therapy, and advances in supportive care. Now that the majority of patients in the western hemisphere reaches adulthood, end organ diseases are frequent, which include vasculopathic complications such as chronic leg ulcers. The management of patients with leg ulcers requires the hematologist to lead a team of health care professionals, and investigates the presence of associated, but potentially still occult signs of vasculopathy, such as pulmonary hypertension, renal disease, priapism and retinopathy. These complications may be asynchronous, and long term careful screening is indicated, in order to ensure early diagnosis and intervention. It is crucial to address both the immediate consequences of pain, infection and disability, and long term effects on quality of life, employment and stigma associated with chronic ulceration. Recent insights into their pathophysiology may have practical implications. We propose a holistic approach to the management of patients' physical and emotional problems and mechanisms of ulcers formation and delayed healing. An overview of topical and systemic therapies for chronic ulcers is given, with the understanding that wound care therapy is best left to the wound specialists, medical and surgical, with whom the hematologist must keep an open line of communication. In the absence of evidence-based guidelines, our opinion is based on both a critical review of the literature and our personal clinical and research experience.

The impact of transcutaneous oxygen pressure measurement in patients with suspected critical lower limb ischemia.

BACKGROUND: Our aims were to describe macro- and microvascular disease by the use of a combination of toe blood pressure (TBP) and transcutaneous oxygen pressure (TcPO2) measurements, and through this approach we aimed to classify lower limb ulcers. One specific aim was to evaluate whether patients with diabetes had a more pronounced disturbance of the microcirculation compared to patients without known diabetes. METHODS: The study included a series of 498 patients with suspected critical lower limb ischemia. All patients were investigated with TBP and TcPO2, and the macro- and microvascular function was graded as normal, reduced or critically reduced. RESULTS: Four hundred and twenty-two patients (85%) had ulcerations at the time of the examination, the same percentage in patients with (254/300) and in patients without diabetes (168/198). Dominantly microvascular disease with TcPO2 more reduced than TBP was seen in 18% (54/300) of the diabetic, and in 17% (33/198) of the non-diabetic patients. In the diabetic patients the lower limb ulcerations were as following: 44% ischemic, 47% mixed and 9% non-ischemic, respectively. In the non-diabetic patients the corresponding numbers were 38%, 52% and 10%. CONCLUSIONS: By combining TBP and TcPO2 the degree of macro- and microvascular dysfunction can be evaluated, classification of lower limb ulcers will be facilitated and the risk of underestimation of the degree of distal ischemia will be reduced. Microvascular dysfunction is as common in non-diabetic patients, as it is in patients with diabetes.

Aging, hypercoagulability, and leg necrosis in critical limb ischemia.

BACKGROUND: Aging is associated with changes in coagulation status and progression of arterial insufficiency. The purpose of this study was to identify interrelationships among aging, coagulation status, and leg necrosis in patients with critical limb ischemia (CLI). METHODS: Between March 2010 and February 2013, 103 consecutive patients with CLI were enrolled in this study. Retrospective analyses were performed on patient characteristics including age, gender, the presence or the absence of leg necrosis, diabetes mellitus (DM), hypertension, and smoking, and preoperatively measured baseline coagulability factors, which included measurements of coagulation factors anticardiolipin antibodies IgG and IgM, lupus anticoagulant and factor 8, the fibrinolytic factor tissue plasminogen activator (t-PA), and natural anticoagulants proteins C and S and antithrombin III. RESULTS: Among 103 patients with CLI, a total of 49 legs from 41 patients presented varying degrees of necrosis. CLI patients with DM and hypertension showed significantly increased incidences of leg necrosis compared with those without (P = 0.000, 0.039, respectively). Patients with CLI and leg necrosis were significantly older compared with the age of those without necrosis (P = 0.007). Blood levels of anticardiolipin antibodies IgG and IgM, factor 8, lupus anticoagulant, and t-PA tended to increase with age. However, blood levels of proteins C and S and antithrombin III decreased with patient age. Patients with CLI and leg necrosis showed significantly increased levels of lupus anticoagulant (P = 0.049) and significantly decreased levels of proteins C and S (P = 0.009 and 0.018, respectively) compared with patients without leg necrosis. CONCLUSIONS: Patients with CLI and leg necrosis were significantly older compared with those without necrosis; similarly, our results revealed age-related hypercoagulability, with significantly elevated coagulation factor lupus anticoagulant and decreased natural anticoagulants protein C and S levels. From these observations, we conclude that age-related hypercoagulability may be an important mechanism that may facilitate leg necrosis in patients with CLI.