Clinical application of fresh fibroblast allografts for the treatment of diabetic foot ulcers: a pilot study

Diabetic foot ulcers often pose a difficult problem for health care professionals because of the defects associated with fibroblast functioning. Although there has been much interest recently in the use of topical growth factors for the treatment of diabetic foot ulcers, the effects are generally not very dramatic. Cryopreserved fibroblast implants, which are able to adjust to a wound's environment and provide the desired growth factors and other substances that may be lacking in a chronic wound, represent an exciting development and a major advance. These products may well provide growth factors in the right concentration and in the right sequence, something that has proved difficult to achieve with the topical application of recombinant growth factors. However, cell activities are impaired by cryopreservation. The purpose of this study was to assess the effects of fresh human allogeneic fibroblast grafting for the treatment of diabetic foot ulcers. Eight patients with diabetic foot ulcers ranging from 6 to 17 weeks in duration were treated. The size of the wounds ranged from 2.0 to 6.0 cm2, with three patients exhibiting exposed bones. A history of diabetic foot ulcers was present in five patients. Human dermal fibroblasts from healthy teenagers were cultured in Dulbecco's modified Eagle medium/Ham's F-12 supplemented with 10% autologous serum. The cultured cells were applied over the wounds immediately after debridement; fibrin was used as a cell carrier. A dressing was then applied with Tegaderm and kept moist until healing was complete. The progress and time for complete wound closure and patient satisfaction were assessed, with follow-up time ranging from 6 to 18 months. Complete wound healing occurred in all patients. Eleven to 21 days were needed for complete reepithelization of the wound, and no clinical or laboratory abnormalities were noted. Patient satisfaction was also very positive. In this study, the use of fresh human fibroblast allografts was found to be a safe and effective treatment for diabetic foot ulcers.     

Single-stage surgical treatment of noninfected diabetic foot ulcers

A retrospective study was undertaken to evaluate a single-stage approach in the treatment of noninfected, chronic, well-perfused diabetic foot wounds. This single-stage approach consisted of total excision of the ulcer with broad exposure, correction of the underlying osseous deformity, and immediate primary closure using a local random flap. Four hundred cases of pedal ulcers were analyzed by chart review. Of those, 67 cases underwent a single-stage surgical treatment and were analyzed for length of hospital stay, postoperative complications, time to heal, recurrence of the ulcer, and postprocedure ambulatory status. The age of the ulcers before surgery was 12 +/- 12 months (mean +/- SD), with a range of 1 to 60. The median perioperative hospital stay was 5 +/- 7.6 days. All patients were followed until the wounds were healed or to amputation. The median total time to heal was 30.8 +/- 40 days. Ninety-seven percent of the wounds healed. The recurrence rate of ulceration was 10.4 percent (seven of 67), over a time span of up to 6 years. All but one patient returned to previous levels of ambulation, and many patients had improved levels of ambulation. The single-stage approach eliminated the need for additional surgical procedures, with their associated costs and risks. In addition, healing times were significantly reduced, resulting in decreased hospital stays and subsequent costs and providing the patient with an expedient return to footwear so that bipedal function could be restored. Most importantly, by addressing the underlying bony pathologic findings, the recurrence rates were also drastically reduced.     

Cellular versus acellular matrix devices in treatment of diabetic foot ulcers: study protocol for a comparative efficacy randomized controlled trial

Background

The referral letter plays a key role both in the communication between primary and secondary care, and in the quality of the health care process. Many studies have attempted to evaluate and improve the quality of these referral letters, but few have assessed the impact of their quality on the health care delivered to each patient.

Methods

A cluster randomized trial, with the general practitioner office as the unit of randomization, has been designed to evaluate the effect of a referral intervention on the quality of health care delivered. Referral templates have been developed covering four diagnostic groups: dyspepsia, suspected colonic malignancy, chest pain, and chronic obstructive pulmonary disease. Of the 14 general practitioner offices primarily served by University Hospital of North Norway Harstad, seven were randomized to the intervention group. The primary outcome is a collated quality indicator score developed for each diagnostic group. Secondary outcomes include: quality of the referral, health process outcome such as waiting times, and adequacy of prioritization. In addition, information on patient satisfaction will be collected using self-report questionnaires. Outcome data will be collected on the individual level and analyzed by random effects linear regression.

Discussion

Poor communication between primary and secondary care can lead to inappropriate investigations and erroneous prioritization. This study’s primary hypothesis is that the use of a referral template in this communication will lead to a measurable increase in the quality of health care delivered.

Trial registration

This trial has been registered at ClinicalTrials.gov. The trial registration number is NCT01470963     

A comparative study of foot dimension between adult male and female and evaluation of foot hazards due to using of footwear

Using footwear often becomes troublesome and creates many problems. Most of these problems are associated with the wearing of ill-fitting footwear, as it leads to biomechanical imbalance and ultimately give rise to different foot problems. In the present investigation different foot problems, viz., discomfort, pain and other hazards related to the use of footwear have been evaluated and attempts have been made to study different foot dimensions of men and women that are related to the design of footwear. For the present study different foot dimensions of both right and left feet of the subjects were measured on 300 Bengalee (Indian) subjects having the age range of 20-35 years. The subjects reported that they had got discomfort, pain, blister and corn due to using different footwear. It was noted that the occurrence of these problems in right foot was greater than that in left foot. There was no significant correlation between foot troubles and type of footwear. Results also showed that there was no significant difference in most of the foot dimensions between left foot and right foot. However, significant difference (P < 0.001) in all foot dimensions was observed between male and female subjects. Correlation coefficient among different foot dimensions has also been evaluated and it was noted that foot length was highly correlated with stature and foot volume, particularly in left foot. Footwear should be made according to the foot dimensions of the user population. The database collected from the Bengalee (Indian) population may be a helpful guide for manufacturing different footwear.     

Tissue therapy with autologous dermal and epidermal culture cells for diabetic foot ulcers

A great part of diabetic ulcers on the lower extremities have difficult healing and represent the most common cause of non-traumatic amputation In case of patients unresponsive to the classical therapy with debridement, dressings and systemic antibiotic therapy, cell therapy may be an excellent indication. The objective of this study was to assess the efficacy of autologous skin cell (fibroblasts and keratinocytes) implants cultivated ex vivo and applied to long-standing (9?C34?years) skin ulcers of five diabetic patients (4 DM2 and 1 DM1) with autologous fibrin glue. There were six ulcers of onset between 4?months and 20?years before and from 4.0 to 36.62?cm² in size, located on the lower limbs and unresponsive to the several conventional treatments. Complete healing was observed in five ulcers (83.3%), after 21?C120?days. The patient who presented the largest ulcer had partial improvement in 40?days. It is believed that the more distal ulcer location is, the worse is its prognosis. There probably is a correlation between healing time, ulcer size and prior duration. No adverse reactions derived from the treatment occurred. It is concluded that this method is an excellent therapeutic option for diabetic ulcers, allowing faster healing. Its great advantage is being a minimally invasive procedure that can be carried out in an outpatient clinic.     

Role of mesenchymal stromal cells derivatives in diabetic foot ulcers: a controlled randomized phase 1/2 clinical trial

Background: Diabetes-related foot complications have been identified as the most common isolated cause of morbidity among patients with diabetes and the leading cause of amputation. Therefore, new strategies to stimulate skin regeneration may provide a novel therapeutic approach to reduce non-healing ulcer disease. Recently, we demonstrated in proof-of-concept in humans that administration of allogeneic bone marrow mesenchymal stromal cellss derivatives (allo-hBM-MSCDs) is effective in a similar way to the use of allogeneic bone marrow mesenchymal stromal cellss (allo-hBM-MSCs) in grade 2 diabetic foot ulcers (DFUs).Aim: To assess the safety and efficacy profile of the allo-hBM-MSCDs relative to the conventional approach (PolyMen® dressing) in 1/2 clinical trial phases in patients with grade 1 and 2 DFUs.Methods: In the present study, we used 2 doses of allo-hBM-MSCDs (1 mL) or 1 dose of allo-hBM-MSCs (1 × 106 cells) intradermally injected around wounds and assessed their safety and effectiveness, relative to the conventional approach (PolyMem dressing). Allo-hBM-MSCDs and allo-hBM-MSCs were produced in a certified Good Manufacturing Practice-type Laboratory. Patients with grade 1 and 2 DFUs were randomized to receive allo-hBM-MSCDs (n=12), allo-hBM-MSCs (n=6) or conventional treatment (PolyMem dressing) (n=10). The wound-healing process was macroscopically evaluated until the complete closure of the ulcers.Results: No adverse events were reported. Patients with grade 1 and 2 DFUs treated with either allo-hBM-MSCDs or allo-hBM-MSCs, achieved greater percentages of wound closure, enhanced skin regeneration in shorter times and a greater ulcer-free survival relative to the patients who received conventional treatment. Finally, through proteomic analysis, we elucidated the proteins and growth factors that are secreted by allo-hBM-MSCs and relevant to the wound-healing process. In addition, by combining proteomics with Gene Ontology analysis, we comprehensively classified secreted proteins on both biological process and molecular function.Conclusions: In this phase 1/2 trial, our cumulative results suggest that 2 doses of allo-hBM-MSCDs combined with a wound dressing are a safe and effective treatment for grade 1 and 2 DFUs.     

Plantar pressures are higher in cases with diabetic foot ulcers compared to controls despite a longer stance phase duration

Background

Current international guidelines advocate achieving at least a 30 % reduction in maximum plantar pressure to reduce the risk of foot ulcers in people with diabetes. However, whether plantar pressures differ in cases with foot ulcers to controls without ulcers is not clear. The aim of this study was to assess if plantar pressures were higher in patients with active plantar diabetic foot ulcers (cases) compared to patients with diabetes without a foot ulcer history (diabetes controls) and people without diabetes or a foot ulcer history (healthy controls).

Methods

Twenty-one cases with diabetic foot ulcers, 69 diabetes controls and 56 healthy controls were recruited for this case-control study. Plantar pressures at ten sites on both feet and stance phase duration were measured using a pre-established protocol. Primary outcomes were mean peak plantar pressure, pressure-time integral and stance phase duration. Non-parametric analyses were used with Holm’s correction to correct for multiple testing. Binary logistic regression models were used to adjust outcomes for age, sex and body mass index. Median differences with 95 % confidence intervals and Cohen’s d values (standardised mean difference) were reported for all significant outcomes.

Results

The majority of ulcers were located on the plantar surface of the hallux and toes. When adjusted for age, sex and body mass index, the mean peak plantar pressure and pressure-time integral of toes and the mid-foot were significantly higher in cases compared to diabetes and healthy controls (p?<?0.05). The stance phase duration was also significantly higher in cases compared to both control groups (p?<?0.05). The main limitations of the study were the small number of cases studied and the inability to adjust analyses for multiple factors.

Conclusions

This study shows that plantar pressures are higher in cases with active diabetic foot ulcers despite having a longer stance phase duration which would be expected to lower plantar pressure. Whether plantar pressure changes can predict ulcer healing should be the focus of future research. These results highlight the importance of offloading feet during active ulceration in addition to before ulceration.

     

Growth hormone exacerbates diabetic renal damage in male but not female rats

Background

Human and animal studies support the idea that there are sex differences in the development of diabetic renal disease. Our lab and others have determined that in addition to Ang II (through the AT₁R), growth hormone (GH) contributes to renal damage in models of renal failure; however, the impact of sex and GH on the mechanisms initiating diabetic renal disease is not known. This study examined the effect of sex and GH on parameters of renal damage in early, uncontrolled streptozotocin (STZ)-induced diabetes.

Methods

Adult male and female Sprague–Dawley rats were injected with vehicle (control), STZ, or STZ?+?GH and euthanized after 8 weeks.

Results

Mild but significant glomerulosclerosis (GS) and tubulointerstitial fibrosis (TIF) was observed in both kidneys from male and female diabetic rats, with GH significantly increasing GS and TIF by 30% and 25% in male rats, but not in female rats. STZ increased TGF-β expression in both kidneys from male and female rats; however, while GH had no further effect on TGF-β protein in diabetic females, GH increased TGF-β protein in the male rat’s kidneys by an additional 30%. This sex-specific increase in renal injury following GH treatment was marked by increased MCP-1 and CD-68+ cell density. STZ also reduced renal MMP-2 and MMP-9 protein expression in both kidneys from male and female rats, but additional decreases were only observed in GH-treated diabetic male rats. The sex differences were independent of AT₁R activity.

Conclusions

These studies indicate that GH affects renal injury in diabetes in a sex-specific manner and is associated with an increase in pro-inflammatory mediators.

     

Variations in the GST activity are associated with single and combinations of GST genotypes in both male and female diabetic patients

In the present cross sectional study, we aimed to ascertain the relative associations of GST genotypes with GST activity variations and also with the risk to DMT2 predisposition among men and women separately. Clinical samples obtained from 244 DMT2 cases (120 Males and 124 Females) and 228 controls (117 Males and 111 Females) belonging to Asian Indian ethnicity were used to test for glycemic index, lipid profile, GST activity and GST genotypes. The frequencies of single and combinations of GST genotypes were statistically examined for their association with DMT2 risk among both study groups. The GST activity is significantly lowered in DMT2 group compared to controls (p = < 0.001). This reduction is found to be subjective to single and combinations of GST genotypes among diabetic patients. The frequency distribution for single, double and triple combinations of genotypes of GSTT1, GSTM1 and GSTP1 showed the varying degrees of association with DMT2 risk from 0.5 to 5.6-fold among male and female patients (for all associations, p value was <0.05). Interestingly, GST activity was lowered in both male and female patients with single or combinational genotypes of GSTM1 (Null), GSTT1 (Null), and P1 (V/V) (for all associations, p value was = <0.0001). The reduced anti-oxidant capacity among diabetic patients with certain GST genotypes may have some important implications for disease diagnosis and therapy.     

Topical hyperbaric oxygen and low energy laser therapy for chronic diabetic foot ulcers resistant to conventional treatment

Chronic foot ulcers are common in long-standing diabetes, may herald severe complications and are often resistant to therapy. To evaluate the effects of adjunctive topical hyperbaric oxygen treatment (THBO) and low energy laser (LEL) irradiation on ulcer healing, a 100 consecutive patients with chronic diabetic foot ulcers (DFU) refractory to 4.5 +/- 1.2 months of comprehensive treatment, were enrolled in a prospective open study. While conventional treatment was continued as necessary, THBO was administered by pumping 100 percent oxygen into a disposable sealed polythylene hyperbaric chamber (150 min x 2 to 3/wk at up to 1.04 atm). Helium-neon LEL irradiation was given concurrently using a Unilaser Scan Unit at 4 J/cm2 for 20 min. Some patients continued THBO at home or their treatment was confined to THBO at home. Patients were monitored every two weeks revealing 81 percent cure after 25 +/- 13 treatments over 3.2 +/- 1.7 months. On follow-up (median 18 months), only 3/81 (4 percent) had reulceration, which responded to THBO/LEL retreatment. Nonresponders had significantly lower ankle brachial indices (ABI) than patients whose ulcers were healed (0.55 vs. 0.78, p < 0.01) and ultimately required amputation. Patient compliance was full and no adverse events occurred. In conclusion, although the study was open and uncontrolled, an 81 percent healing of DFU in patients who previously did not respond to a comprehensive treatment program, constitutes an intriguing preliminary result. Thus, THBO/LEL therapy may be a safe, simple, and inexpensive early adjunctive treatment for patients with chronic diabetic foot ulcers. Our findings should prompt its evaluation by large randomized controlled trials.     

Patterns of reproductive effort in male and female shrubs of Oemleria cerasiformis: a 6-year study

1 We monitored flowering and fruiting of individual male and female plants of Oemleria cerasiformis over a 6-year period in a population in western Canada, and calculated fruit set (percentage of pistils maturing) and reproductive effort (RE) (gram of reproductive tissue per gram of leaf).
2 Over 6 years, male O. cerasiformis had on average much lower total RE, but much higher RE at flowering, than females.
3 In males, strong correlations between RE and light suggested that investment in reproduction was largely determined by light levels. There were strong positive correlations of RE between years, with no evidence of periodic fluctuations.
4 In females, in contrast to males, RE at flowering was not related to light. However, fruit set was strongly correlated with light. Flowering RE and fruit set were uncorrelated in females, indicating that these are affected by different factors.
5 Correlations of RE between years in females, although often significant, were lower than in males, indicating that RE fluctuates more between years in females than in males and may respond to past levels of RE. Flowering may reflect adjustments in response to past reproduction, or may be controlled by resources other than light. Fruit set was not significantly related to previous RE.
6 The greater total RE of females and their limited ability to adjust fruit set are probably major factors contributing to the greater mortality rates of females and the male-biased sex ratios in O. cerasiformis .     

Polyherbal formulation exerts wound healing,anti-inflammatory,angiogenic and antimicrobial properties: Potential role in the treatment of diabetic foot ulcers

Diabetic foot ulcer (DFU) is a common and devastating complication in diabetic patients and is associated with an elevated risk of amputation and mortality. DFU remains a major therapeutic challenge due to poor understanding of its underlying pathogenesis. This complication is characterized by impaired wound healing; however, mechanisms causing this impairment are complicated and involve interactions between many different cell types and infections. In addition to other conventional DFU treatments, herbal foot baths are also common, although little is known about their mechanisms of action, and they contain a wide variety of herbal ingredients. In this study, we aimed to examine the effects of three polyherbal formulations consisting of medicinal plants used in traditional Thai herbal foot baths on wound healing, anti-inflammation, angiogenesis, and extracellular matrix modulation using high-concentration glucose-treated human keratinocytes, in addition to antibacterial evaluation. Our results showed that formulation 3 (F3) possessed the greatest potential to restore the impairment of keratinocytes caused by high glucose concentrations. We found that F3 could inhibit the growth of Staphylococcus aureus, accelerate wound healing, and upregulate the expression of TIMP-1, VEGF, and TGF-β, and downregulate the expression of TNF-α, IL-6, and MMP-9. Collectively, these data support the potential of F3 for therapeutic development in the treatment of DFU.     

Nest ornamentation by female spotless starlings in response to a male display: an experimental study

1. The use of behavioural traits by females in signalling condition has been practically ignored in evolutionary theory. However, females may also exhibit ornaments and behavioural displays, although less elaborated than those of males. 2. In this study we suggest that the carrying of feathers by spotless starlings Sturnus unicolor Temminck females to decorate the nest represents an elaborated and costly behaviour that is displayed in response to a courtship male behaviour: the carrying of nest green plants. 3. By experimentally increasing the amount of green plants in the nests, to give the appearance that highly attractive males defended them, we induced females to increase their feather carrying rates. 4. The amount of feathers carried to the nest was correlated to female reproductive experience and laying date, two variables correlated with female body condition. These results suggests that this behaviour may work as an honest indicator of female quality. 5. We conclude that male carrying plants and female carrying feathers can be viewed as two sex-specific functionally related signalling behaviours involved in mutual courtship or status signalling.     

Recurrent peptic ulcers in patients following successful Helicobacter pylori eradication: a multicenter study of 4940 patients

Objective. Although curative treatment of Helicobacter pylori infection markedly reduces the relapse of peptic ulcers, the details of the ulcers that do recur is not well characterized. The aim of this study is to describe the recurrence rate and specific features of peptic ulcers after cure of H. pylori infection. Methods. This was a multicenter study involving 4940 peptic ulcer patients who were H. pylori negative after successful eradication treatment and were followed for up to 48 months. The annual incidence of ulcer relapse in H. pylori‐cured patients, background of patients with relapsed ulcers, time to relapse, ulcer size, and site of relapsed ulcers were investigated. Results. Crude peptic ulcer recurrence rate was 3.02% (149/4940). The annual recurrence rates of gastric, duodenal and gastroduodenal ulcer were 2.3%, 1.6%, and 1.6%, respectively. Exclusion of patients who took NSAIDs led annual recurrence rates to 1.9%, 1.5% and 1.3%, respectively. The recurrence rate was significantly higher in gastric ulcer. Recurrence rates of patients who smoked, consumed alcohol, and used NSAIDs were significantly higher in those with gastric ulcer recurrence compared to duodenal ulcer recurrence (e.g. 125 of 149 [83.9%] relapsed ulcers recurred at the same or adjacent sites as the previous ulcers). Conclusions. Curative treatment of H. pylori infection is useful in preventing ulcer recurrence. Gastric ulcer is more likely to relapse than duodenal ulcer. Recurrent ulcer tended to recur at the site of the original ulcers.     

Overexpression of the gap junction protein Cx43 as found in diabetic foot ulcers can retard fibroblast migration

Poor healing of DFUs (diabetic foot ulcers) is a major clinical problem that can be extremely debilitating and lead to lower limb amputation. In the normal acute wound, the Cx43 (connexin 43) gap junction protein is down-regulated at the wound edge as a precursor to cell migration and healing. In fibroblasts from the human chronic DFU wound edge there was a striking and significant 10-fold elevation of Cx43 protein, as well as a 6-fold increase in N-cadherin and a 2-fold increase in ZO-1 (zonular occludin-1), compared with unwounded skin. In streptozotocin diabetic rats, Cx43 was found to be up-regulated in intact dermal fibroblasts in direct proportion to blood glucose levels and increased 2-fold further in response to wounding of the skin. To mimic diabetes, NIH 3T3 fibroblasts were cultured under different concentrations of glucose or mannitol and Cx43 protein intercellular communication and migration rates were determined. Cultures of fibroblasts in very high (40 mM) glucose conditions showed significantly elevated Cx43 protein levels, as shown by immunostaining and Western blotting, and significantly increasing gap junctional communication, as shown by dye transfer. In scratch wound-healing assays, increased levels of Cx43 from high glucose resulted in repressed filopodial extensions and significantly slower migration rates than in either standard conditions (5.5 mM glucose) or the osmotic control of mannitol. Conversely, when glucose-induced Cx43 up-regulation was prevented with Cx43shRNA (Cx43 short-hairpin RNA) transduction, the fibroblasts extended long filopodia and migrated significantly faster. Cx43 protein was up-regulated in fibroblasts in DFUs as well as after high glucose exposure in culture which correlated with inhibition of fibroblast migration and is likely to contribute to impaired wound healing.