Pedro Lopez-Saura

Methylglyoxal administration induces diabetes-like microvascular changes and perturbs the healing process of cutaneous wounds.

Increased formation of MG (methylglyoxal) and related protein glycation in diabetes has been linked to the development of diabetic vascular complications. Diabetes is also associated with impaired wound healing. In the present study, we investigated if prolonged exposure of rats to MG (50-75 mg/kg of body weight) induced impairment of wound healing and diabetes-like vascular damage. MG treatment arrested growth, increased serum creatinine, induced hypercholesterolaemia (all P < 0.05) and impaired vasodilation (P < 0.01) compared with saline controls. Degenerative changes in cutaneous microvessels with loss of endothelial cells, basement membrane thickening and luminal occlusion were also detected. Acute granulation appeared immature (P < 0.01) and was associated with an impaired infiltration of regenerative cells with reduced proliferative rates (P < 0.01). Immunohistochemical staining indicated the presence of AGEs (advanced glycation end-products) in vascular structures, cutaneous tissue and peripheral nerve fibres. Expression of RAGE (receptor for AGEs) appeared to be increased in the cutaneous vasculature. There were also pro-inflammatory and profibrotic responses, including increased IL-1beta (interleukin-1beta) expression in intact epidermis, TNF-alpha (tumour necrosis factor-alpha) in regions of angiogenesis, CTGF (connective tissue growth factor) in medial layers of arteries, and TGF-beta (transforming growth factor-beta) in glomerular tufts, tubular epithelial cells and interstitial endothelial cells. We conclude that exposure to increased MG in vivo is associated with the onset of microvascular damage and other diabetes-like complications within a normoglycaemic context.

The pro-inflammatory environment in recalcitrant diabetic foot wounds

Lower extremity ulceration is one of the serious and long‐term diabetic complications rendering a significant social burden in terms of amputation and quality‐of‐life reduction. Diabetic patients experience a substantial wound‐healing deficit. These lesions are featured by an exaggerated and prolonged inflammatory reaction with a significant impairment in local bacterial invasion control. Experimental and clinical evidences document the deleterious consequences of the wound’s pro‐inflammatory phenotype for the repair process. From a biochemical standpoint, hyperinflammation favours wound matrix degradation, thus, amplifying a pre‐existing granulation tissue productive cells’ invasiveness and recruitment deficit. Tumour necrosis factor perpetuates homing of inflammatory cells, triggers pro‐apoptotic genes and impairs reepithelialisation. Advanced glycation end‐products act in concert with inflammatory mediators and commit fibroblasts and vascular cells to apoptosis, contributing to granulation tissue demise. Therapeutic approaches aimed to downregulate hyperinflammation and/or attenuate glucolipotoxicity may assist in diabetic wound healing by dismantling downstream effectors. These medical interventions are demanded to reduce amputations in an expanding diabetic population.

Thrombolysis as first choice therapy in prosthetic heart valve thrombosis. A study of 68 patients.

AbstractBackground and objectives: Valvular thrombosis is a serious complication in patients with prosthetic heart valves. Traditional treatment is emergency surgery, but thrombolysis provides a non invasive alternative. In this paper we evaluate the efficacy and safety of thrombolysis in prosthetic heart valve thrombosis. Methods: Data of 68 patients diagnosed of prosthetic valve thrombosis, treated at the Institute of Cardiology and Cardiovascular Surgery, Havana during a 6-years period were analyzed. They received thrombolysis with a recombinant streptokinase infusion at 250 000 IU in 30 minutes followed by 100 000 IU/hour during 72 hours or less if the thrombosis resolved before. The evaluation was based on clinical and echocardiographic findings. Results: Affected sites were mitral (50 cases), tricuspid (9), and aortic (9). Mean time of prosthesis implantation was 6.8 years. The presentation form was generally heart failure (NYHA functional class III–IV) in 64 (94.1%) patients. Mean time interval between onset of symptoms and diagnosis was 10.6 days. There was total response to treatment in 58 (85.3%) patients, partial in 4 (5.9%) and failure in 6 (8.8%). Recombinant streptokinase overall dose was 5.1 × 106 IU and mean infusion time 50 hours. Major hemorrhagic complications were observed in two patients. Five embolic events occurred during thrombolysis. Four patients died. Rethrombosis was noted in 11 patients; 10 were retreated successfully with thrombolysis. Conclusions: Thrombolysis with recombinant streptokinase is efficacious and safe for the treatment of prosthetic heart valve thrombosis. It does not contraindicate surgical treatment if there is no total response, because patient goes to surgery in better hemodynamic conditions with lower risk. Nowadays it can be considered as first-line treatment in all patients with prosthetic heart valve thrombosis regardless of functional class unless specific contraindications exist.

Intra-lesional injections of recombinant human epidermal growth factor promote granulation and healing in advanced diabetic foot ulcers: multicenter, randomised, placebo-controlled, double-blind study

Fernández‐Montequín JI, Valenzuela‐Silva CM, González Díaz O, Savigne W, Sancho‐Soutelo N, Rivero‐Fernández F, Sánchez‐Penton P, Morejón‐Vega L, Artaza‐Sanz H, García‐Herrera A, González‐Benavides C, Hernández‐Cañete CM, Vázquez‐Proenza A, Berlanga‐Acosta J, López‐Saura PA, for the Cuban Diabetic Foot Study Group. Intra‐lesional injections of recombinant human epidermal growth factor promote granulation and healing in advanced diabetic foot ulcers: multicenter, randomised, placebo‐controlled, double‐blind study.

Epidermal growth factor in clinical practice – a review of its biological actions, clinical indications and safety implications

Chemotaxis, mitogenesis, motogenesis and cytoprotection are common cellular events involved in both tumourigenesis and tissue repair, which appear amplified upon growth factors exposure. Epidermal growth factor (EGF) promotes these events in epithelial and mesenchymal cells through the binding to a specific tyrosine kinase receptor. In experimental oncology settings, EGF does not initiate malignant transformation but exhibits ‘tumour promotion’. These observations have raised doubts on the clinical use of EGF despite solid demonstrations of efficacy in experimental conditions and clinical trials. The results of a Pubmed and Bioline investigation on EGF clinical uses and preclinical safety data are presented here. EGF topical administration has been used since 1989 to enhance the healing process of a variety of peripheral tissues wounds (16 clinical reports), as well as its intravenous, oral and rectal administration for gastrointestinal damages (11 clinical reports). EGF therapeutic efficacy and excellent tolerability seem demonstrated. Lack of long‐term adverse effects is highlighted in those studies with 6, 12 and 24 months of patients follow‐up. Although post‐treatment follow‐up may fall short for malignant growth, there are no reports on evidences linking EGF clinical use with cancer. A multicentre, nationwide survey in Cuba, 15 years after randomly using silver sulphadiazine with EGF or not in burn victims yielded that cancer incidence was comparable between EGF‐treated and control subjects and that such incidence rate does not differ from the age‐matched national incidence for those 15‐year period. All the animal species subjected to long‐term EGF systemic administration exhibit dose‐dependent and reversible epithelial organs hyperplasia with no changes in cells phenotypic differentiation. Histotypic pre‐malignant markers were not identified. The results emerged from co‐carcinogenesis studies and from transgenic mice over‐expressing EGF are conflicting and indicate that EGF overexposure, either innate or postnatal, may not be sufficient to transform cells. The ability of EGF to heal injured tissues in life‐threatening scenarios or to assist in preventing physical and social disability advocates for its clinical use under a rational medical risk/benefit balance.

Adjuvant interferon gamma in patients with drug – resistant pulmonary tuberculosis: a pilot study

BackgroundTuberculosis (TB) is increasing in the world and drug-resistant (DR) disease beckons new treatments.MethodsTo evaluate the action of interferon (IFN) gamma as immunoadjuvant to chemotherapy on pulmonary DR-TB patients, a pilot, open label clinical trial was carried out in the Cuban reference ward for the management of this disease. The eight subjects existing in the country at the moment received, as in-patients, 1 × 106 IU of recombinant human IFN gamma intramuscularly, daily for one month and then three times per week up to 6 months as adjuvant to the indicated chemotherapy, according to their antibiograms and WHO guidelines. Sputum samples collection for direct smear observation and culture as well as routine clinical and thorax radiography assessments were done monthly.ResultsSputum smears and cultures became negative for acid-fast-bacilli before three months of treatment in all patients. Lesion size was reduced at the end of 6 months treatment; the lesions disappeared in one case. Clinical improvement was also evident; body mass index increased in general. Interferon gamma was well tolerated. Few adverse events were registered, mostly mild; fever and arthralgias prevailed.ConclusionsThese data suggest that IFN gamma is useful and well tolerated as adjunctive therapy in patients with DR-TB. Further controlled clinical trials are encouraged.

Intralesional injections of Citoprot‐P® (recombinant human epidermal growth factor) in advanced diabetic foot ulcers with risk of amputation

To investigate the efficacy and safety of recombinant human epidermal growth factor (rhEGF) in advanced diabetic foot ulcers (DFU) A double‐blind trial was carried out to test two rhEGF dose levels in type 1 or 2 diabetes patients with Wagner’s grade 3 or 4 ulcers, with high risk of amputation. Subjects were randomised to receive 75 (group I) or 25 μg (group II) rhEGF through intralesional injections, three times per week for 5–8 weeks together with standardised good wound care. Endpoints were granulation tissue formation, complete healing and need of amputation. Safety was assessed by clinical adverse events (AEs) and laboratory evaluations. Forty‐one patients were included. After 5–8 weeks of treatment, 83% patients in the higher dose group and 61% in group II achieved useful granulation tissue covering more than 98% of the wound area. At long‐term assessment, 13 (56·5%) patients healed in group I and 9 (50%) in group II. The mean time to complete healing in group I was 20·6 weeks (95% CI: 17·0–24·2) and 19·5 weeks (16·3–22·7) in group II. After 1‐year follow‐up, only one patient relapsed. Amputation was not necessary in 65% and 66·7% of groups I and II, respectively. The AEs rates were similar. The most frequent were sepsis (33%), burning sensation (29%), tremors, chills and local pain (25% each). rhEGF local injection enhances advanced DFU healing and reduces the risk of major amputation. No dose dependency was observed.

Epidermal growth factor intralesional infiltrations can prevent amputation in patients with advanced diabetic foot wounds

This study examined if a series of epidermal growth factor (EGF) local infiltrations can enhance the healing process of complicated diabetic wounds. Twenty‐nine in‐hospital patients with diabetic neuropathic or ischaemic lesions with high risk of amputation were treated in a non controlled pilot study conducted at the National Institute of Angiology, Havana. Lesions, classified as Wagner’s grade 3 or 4, included ulcers ≥20 cm2 for ≥25 days or amputation residual bases ≥30 cm2 for ≥15 days, healing refractory despite comprehensive wound care. EGF (25 μg) intralesional infiltrations (≈250 μl of a 25 μg/ml solution/injection point) were performed thrice weekly up to the eighth week. Wound closure was monitored during the treatment and recurrence examined for a year following discharge from hospital. Eighty‐six per cent of the patients treated showed a productive granulation at infiltration session 8. Histological examination at this point indicated a substantial wound matrix transformation, granulation tissue cell repopulation and angiogenesis. Of the 29 patients treated, amputation was prevented in 17 (58·6%) of them who completed 24 infiltration sessions. They averaged 71·1 ± 18·3% of reepithelisation during a mean in‐hospital period of 66·5 ± 4·9 days. Wound recurrence after 1 year of follow‐up appeared in only one patient. Preliminary evidences suggest that EGF intralesional infiltrations may be effective in reducing diabetic lower limb amputation.

C-Phycocyanin is neuroprotective against global cerebral ischemia/reperfusion injury in gerbils

Although the huge economic and social impact and the predicted incidence increase, neuroprotection for ischemic stroke remains as a therapeutically empty niche. In the present study, we investigated the rationale of the C-Phycocyanin (C-PC) treatment on global cerebral ischemia/reperfusion (I/R) injury in gerbils. We demonstrated that C-PC given either prophylactically or therapeutically was able to significantly reduce the infarct volume as assessed by triphenyltetrazolium chloride (TTC) staining and the neurological deficit score 24h post-stroke. In addition, C-PC exhibited a protective effect against hippocampus neuronal cell death, and significantly improved the functional outcome (locomotor behavior) and gerbil survival after 7 days of reperfusion. Malondialdehyde (MDA), peroxidation potential (PP) and ferric reducing ability of plasma (FRAP) were assayed in serum and brain homogenates to evaluate the redox status 24h post-stroke. The treatment with C-PC prevented the lipid peroxidation and the increase of FRAP in both tissue compartments. These results suggest that the protective effects of C-PC are most likely due to its antioxidant activity, although its anti-inflammatory and immuno-modulatory properties reported elsewhere could also contribute to neuroprotection. To our knowledge, this is the first report of the neuroprotective effect of C-PC in an experimental model of global cerebral I/R damage, and strongly indicates that C-PC may represent a potential preventive and acute disease modifying pharmacological agent for stroke therapy.

Adjuvant interferon gamma in patients with pulmonary atypical Mycobacteriosis: A randomized, double-blind, placebo-controlled study

BackgroundHigh antibiotic resistance is described in atypical Mycobacteriosis, mainly by Mycobacterium avium complex (MAC).MethodsA randomized, double-blind, placebo-controlled clinical trial was carried out in two hospitals to evaluate the effect of interferon (IFN) gamma as immunoadjuvant to chemotherapy on patients with atypical mycobacteria lung disease. Patients received placebo or 1 × 106 IU recombinant human IFN gamma intramuscularly, daily for one month and then three times per week up to 6 months as adjuvant to daily oral azithromycin, ciprofloxacin, ethambutol and rifampin. Sputum samples collection for direct smear observation and culture as well as clinical and thorax radiography assessments were done during treatment and one year after. Cytokines and oxidative stress determinations were carried out in peripheral blood before and after treatment.ResultsEighteen patients were included in the IFN group and 14 received placebo. Groups were homogeneous at entry; average age was 60 years, 75% men, 84% white; MAC infection prevailed (94%). At the end of treatment, 72% of patients treated with IFN gamma were evaluated as complete responders, but only 36% in the placebo group. The difference was maintained during follow-up. A more rapid complete response was obtained in the IFN group (5 months before), with a significantly earlier improvement in respiratory symptoms and pulmonary lesions reduction. Disease-related deaths were 35.7% of the patients in the placebo group and only 11.1% in the IFN group. Three patients in the IFN group normalized their globular sedimentation rate values. Although differences in bacteriology were not significant during the treatment period, some patients in the placebo group converted again to positive during follow-up. Significant increments in serum TGF-beta and advanced oxidation protein products were observed in the placebo group but not among IFN receiving patients. Treatments were well tolerated. Flu-like symptoms predominated in the IFN gamma group. No severe events were recorded.ConclusionThese data suggest that IFN gamma is useful and well tolerated as adjuvant therapy in patients with pulmonary atypical Mycobacteriosis, predominantly MAC. Further wider clinical trials are encouraged.Trial registrationCurrent Controlled Trials ISRCTN70900209.