Katarina Fagher

Decreased phosphatase activity, increased Ca2+ sensitivity, and myosin light chain phosphorylation in urinary bladder smooth muscle of newborn mice

Developmental changes in the regulation of smooth muscle contraction were examined in urinary bladder smooth muscle from mice. Maximal active stress was lower in newborn tissue compared with adult, and it was correlated with a lower content of actin and myosin. Sensitivity to extracellular Ca2+ during high-K+ contraction, was higher in newborn compared with 3-wk-old and adult bladder strips. Concentrations at half maximal tension (EC50) were 0.57 ± 0.01, 1.14 ± 0.12, and 1.31 ± 0.08 mM. Force of the newborn tissue was inhibited by ∼45% by the nonmuscle myosin inhibitor Blebbistatin, whereas adult tissue was not affected. The calcium sensitivity in newborn tissue was not affected by Blebbistatin, suggesting that nonmuscle myosin is not a primary cause for increased calcium sensitivity. The relation between intracellular [Ca2+] and force was shifted toward lower [Ca2+] in the newborn bladders. This increased Ca2+ sensitivity was also found in permeabilized muscles (EC50: 6.10 ± 0.07, 5.77 ± 0.08, and 5.55 ± 0.02 pCa units, in newborn, 3-wk-old, and adult tissues). It was associated with an increased myosin light chain phosphorylation and a decreased rate of dephosphorylation. No difference was observed in the myosin light chain phosphorylation rate, whereas the rate of myosin light chain phosphatase–induced relaxation was about twofold slower in the newborn tissue. The decreased rate was associated with a lower expression of the phosphatase regulatory subunit MYPT-1 in newborn tissue. The results show that myosin light chain phosphatase activity can be developmentally regulated in mammalian urinary bladders. The resultant alterations in Ca2+ sensitivity may be of importance for the nervous and myogenic control of the newborn bladders.

Decreased shortening velocity and altered myosin isoforms in guinea‐pig hypertrophic intestinal smooth muscle

The aims of this study were to investigate whether hypertrophy of the small intestinal smooth muscle leads to alterations of myosin isoform composition and shortening velocity and whether possible changes correlate with a change in the sensitivity to ADP of shortening velocity in this tissue. A partial occlusion was introduced in the distal part of the ileum of guinea‐pigs. After 2 weeks, the part of the small intestine just proximal of the stenosis was hypertrophied (indicated by a significantly increased cross‐sectional area). The most proximal part of the small intestine was used as control, thus enabling comparisons between hypertrophic and normal tissue from the same animal. The outer longitudinal layer of the intestinal wall was gently peeled off and used for biochemistry, RT‐PCR and mechanical experiments. The desmin/actin ratio was significantly increased following hypertrophy, although myosin and actin expression were similar in control and hypertrophic tissue. In hypertrophic tissue, the myosin heavy chain mRNA with a 21 base pair insert decreased significantly. The composition of the mRNA encoding the myosin essential light chains changed towards more of the basic type (LC17b). No change in the expression of non‐muscle myosin heavy chains A and B was detected. The maximal shortening velocity (Vmax) of maximally activated skinned preparations was significantly lower in the hypertrophic tissue (≈50 % of control). The sensitivity of Vmax to ADP was increased in the hypertrophic smooth muscle tissue. We conclude that myosin expression is altered following intestinal hypertrophy and that these alterations affect reactions in the cross‐bridge interaction, leading to a slower and more economical contractile function.

What is the Role of Hyperbaric Oxygen in the Management of Diabetic Foot Disease

Systemic hyperbaric oxygen (HBO) is accomplished when a patient is breathing 100% oxygen in an environment with increased barometric pressure. A typical HBO treatment protocol of diabetic foot ulcer involves 20 to 40 sessions. Treatment is usually given as daily 90- to 120-minute HBO sessions at pressures between 2.0 and 2.5 absolute atmospheres. The wide use of HBO as treatment of diabetic foot ulcers over the past decades has been founded on weak scientific ground (ie, few and small prospective studies with methodologic limitations on top of case series). However, the consistency in positive outcome in these trials evaluating HBO on ulcer healing is noteworthy because these findings are in concert with data from in vitro and physiologic studies supporting the theoretic framework of HBO reversing hypoxia-induced pathology. Two well-designed randomized double-blinded placebo-controlled studies have in recent years put HBO on firmer ground as treatment of a selection of diabetic patients with chronic foot. Some evidence indicates that microvascular parameters such as transcutaneous (partial) oxygen pressure (TcPO2) could be useful in predicting which patients will benefit from therapy. Health economic studies suggest potential cost-effectiveness of HBO. But because these analyses are limited by their deficient primary clinical data, they should be interpreted with caution. Thus, HBO is only indicated in a selected group of patients with chronic diabetic foot ulcers. Several key issues remain to be addressed such as developing robust criteria to determine which patients are likely to benefit and when to start and stop treatment.

Effects of thyroxine on myosin isoform expression and mechanical properties in guinea-pig smooth muscle.

Information on the effects of thyroid hormone on smooth muscle contractile protein expression and mechanical properties is sparse. We have addressed the following questions. (1) Can thyroxine hormone alter myosin isoform composition in smooth muscle? (2) Can a change in myosin isoform composition lead to altered mechanical properties in smooth muscle? (3) Are alterations, if occurring, equal in fast and slow smooth muscle types? Guinea‐pigs were treated with thyroxine (T4) for 12 days. Control animals were given physiological saline solution. Maximal unloaded shortening velocity (Vmax) was measured in chemically skinned, maximally activated muscle preparations from the aorta and the taenia coli. Vmax increased following thyroxine treatment, by approximately 20 % in the taenia coli. In the aorta, no significant increase in Vmax could be detected. The sensitivity of isometric force to inorganic phosphate (Pi) was increased in the taenia coli following thyroxine treatment. The expression of mRNA (determined with RT‐PCR) for the myosin heavy chain with the seven amino acid insert increased by approximately 70 % in the aorta and about 25 % in the taenia coli following thyroxine treatment. Western blot analysis showed an increase in the inserted myosin heavy chain form in the taenia coli. Expression of mRNA for the myosin essential light chains and the corresponding proteins did not change significantly in either muscle type. No alterations in non‐muscle myosin heavy chain isoforms could be detected after thyroxine treatment. In conclusion, thyroxine treatment alters the isoform composition of myosin in fast and slow smooth muscles in vivo. This change is sufficient to increase shortening velocity and sensitivity of isometric force to Pi in the fast, but not in the slow, smooth muscle type.

Heart rate-corrected QT interval prolongation as a prognostic marker for 3-year survival in people with Type 2 diabetes undergoing above-ankle amputation.

To evaluate whether heart rate‐corrected QT interval is a plausible prognostic factor for survival after major amputation in people with Type 2 diabetes.

Hyperbaric oxygen therapy reduces the risk of QTc interval prolongation in patients with diabetes and hard-to-heal foot ulcers

AIMS Heart rate corrected QT (QTc) interval prolongation is a risk factor associated with increased mortality. Hyperbaric oxygen therapy (HBO) has previously been shown to have acute beneficial effects on QTc dispersion. The aim of this study was to evaluate long-term effects of HBO on QTc time in diabetic patients with hard-to-heal foot ulcers. METHODS In a prospective, double-blinded placebo-controlled study, patients were randomized to 40 treatment sessions with either HBO or air (placebo), at 2.5 ATA. Patients fulfilling >35 completed treatment sessions were included in the evaluation. RESULTS Of the initial 75 patients (38 HBO/37 placebo), two were excluded due to pacemaker use. Baseline characteristics were similar between groups. At the 2-year follow-up, QTc time was significantly shorter in the HBO compared to the placebo group (438 vs. 453ms, p<0.05). Further, fewer HBO treated patients had a QTc time >450ms (22 vs. 53 %, p<0.02). This difference seemed to be caused by a significant prolongation of the QTc interval in the placebo group (427 (419-459) at baseline vs. 456ms (424-469) after 2years), whereas no significant change was seen in HBO treated patients. CONCLUSIONS HBO treatment might protect against QTc prolongation in this high-risk diabetic population.

Comment on Fedorko et al. Hyperbaric oxygen therapy does not reduce indications for amputation in patients with diabetes with nonhealing ulcers of the lower limb : A prospective, double-blind, randomized controlled clinical trial. Diabetes care 2016; 39:392-399

More than 3 years after the last study visit, the study by Fedorko et al. (1) was published in Diabetes Care . Several issues need to be highlighted, some of which are discussed here. If the end point “indication for [major] amputation” is more appropriate than “major amputation” (1), only the future can tell. Amputation rate assumption of 39.39 vs. 11.54% within a 3-month period in an outpatient care setting seems odd. Accordingly, amputation rates in the trial by Faglia et al. including hospitalized patients with severe and acute infected Wagner grade 3/4 ulcers were 9 vs. 33%, and the 1-year major amputation rate in the outpatient care studies by Abidia …