Serum levels of glucose, thyroid stimulating hormone, and free thyroxine in boys diagnosed with attention deficit hyperactivity disorder: a cross-sectional pilot study.

BACKGROUND: Although attention deficit hyperactivity disorder (ADHD) is a common neurodevelopmental disorder, its aetiology remains unclear. We aimed to establish a relationship between ADHD diagnosis and serum levels of glucose, free thyroxine (FT4), and thyroid stimulating hormone (TSH) in primary school aged boys. METHODS: In a cross-sectional study, we enrolled 133 participants aged 6.5-12.5 years, 67 of whom met DSM-5 criteria for ADHD and 66 healthy age-matched boys. The ADHDT test (ADHDT) was used to assess ADHD symptoms and the Wechsler Intelligence Scale for Children - Revised was used to exclude participants with cognitive deficits. The ADHD participants were tested using the Iowa Conners' Teacher Rating Scale. RESULTS: The ADHD participants had lower glucose levels, higher TSH values, and significantly lower FT4 values than the control group. The multiple logistic regression analysis showed that TSH is a parameter that is 2.7% more likely to occur in the ADHD group. We found a significant correlation between the TSH level and the symptoms of hyperactivity (r = 0.318, p = 0.009) and impulsivity (r = 0.275, p = 0.024) as well as between the glucose level and the symptoms of hyperactivity (r = 0.312, p = 0.010). CONCLUSIONS: Certain ADHD symptoms may correlate with certain hormonal patterns. Our results suggest that the likelihood of suffering from ADHD was lower when FT4 levels were elevated. One biochemical parameter that was significantly and independently associated with the diagnosis of ADHD was the serum TSH level. TRIAL REGISTRATION: On June 26, 2018, at its VI session in 2018, the Ethics Committee of the Institute for Mental Health in Belgrade, Serbia, has considered and unanimously approved the conduct of the research, under the number 1704/1.

Computerized 4-choice reaction time test for the measurement of psychomotor recovery after general anesthesia.

The recovery of patients after general anesthesia is usually estimated by using clinical scores. Since there is a lack of objective methods for assessing psychomotor recovery, the aim of this study was to evaluate three psychological tests for this purpose. Patients, scheduled for ambulatory gynecological surgery, underwent 3 standard psychological tests before (T1), 15 min after the surgery (T2) and on discharge from the recovery room (T3). The tests used were Wechsler memory scale (test 1, working memory capacity), d2-test (test 2, concentration endurance) and computer-based 4-choice-reaction time (4CRT, test 3, reaction time) as well as Postanesthesia Discharge Scoring System (PADSS). The same test battery was used in healthy female volunteers, all test results were compared at the different time points. In 109 patients, working memory capacity and concentration (tests 1 and 2) decreased, the reaction time (test 3) was prolonged at T2 in comparison with T1 and T3 (P < 0.01). PADSS increased from 8 (T2) to 10 (T3) (medians, P < 0.001). Fifty-seven healthy volunteers demonstrated a practice effect in all 3 tests through the course of the study (P <0.01). 4CRT test had shortest duration and enabled computerized data processing. All three tests objectively assess the recovery of psychomotor function in patients after general anesthesia, the computer-based 4CRT seems to be the most convenient for the clinical routine.

Traumatic brain injury: neuropathological, neurocognitive and neurobehavioral sequelae.

Traumatic brain injury (TBI) causes substantial neurological disabilities and mental distress. Annual TBI incidence is in magnitude of millions, making it a global health challenge. Categorization of TBI into severe, moderate and mild by scores on the Glasgow coma scale (GCS) is based on clinical grounds and standard brain imaging (CT). Recent research focused on repeated mild TBI (sport and non-sport concussions) suggests that a considerable number of patients have long-term disabling neurocognitive and neurobehavioral sequelae. These relate to subtle neuronal injury (diffuse axonal injury) visible only by using advanced neuroimaging distinguishing microstructural tissue damage. With advanced MRI protocols better characterization of TBI is achievable. Diffusion tensor imaging (DTI) visualizes white matter pathology, susceptibility weight imaging (SWI) detects microscopic bleeding while functional magnetic resonance imaging (fMRI) provides closer understanding of cognitive disorders etc. However, advanced imaging is still not integrated in the clinical care of patients with TBI. Patients with chronic TBI may experience many somatic disorders, cognitive disturbances and mental complaints. The underlying pathophysiological mechanisms occurring in TBI are complex, brain injuries are highly heterogeneous and include neuroendocrine dysfunctions. Post-traumatic neuroendocrine dysfunctions received attention since the year 2000. Occurrence of TBI-related hypopituitarism does not correlate to severity of the GCS scores. Complete or partial hypopituitarism (isolated growth hormone (GH) deficiency as most frequent) may occur after mild TBI equally as after moderate-to-severe TBI. Many symptoms of hypopituitarism overlap with symptoms occurring in patients with chronic TBI, i.e. they have lower scores on neuropsychological examinations (cognitive disability) and have more symptoms of mental distress (depression and fatigue). The great challenges for the endocrinologist are: (1) detection of hypopituitarism in patients with TBI prospectively (in the acute phase and months to years after TBI), (2) assessment of the extent of cognitive impairment at baseline, and (3) monitoring of treatment effects (alteration of cognitive functioning and mental distress with hormone replacement therapy). Only few studies recently suggest that with growth hormone (rhGH) replacement in patients with chronic TBI and with abnormal GH secretion, cognitive performance may not change while symptoms related to depression and fatigue improve. Stagnation in post-TBI rehabilitation progress is recommended as a signal for clinical suspicion of neuroendocrine dysfunction. This remains a challenging area for more research.

Impact of lipid modulation on the intestinal microcirculation in experimental sepsis.

It has been observed, that patients who were treated medically for dyslipoproteinemia had a potentially lower risk of complications during infection and sepsis, regarding both morbidity and mortality. Aim of this study in experimental sepsis was to elucidate the impact of lipid metabolism modulation by simvastatin, HDL, or bezafibrate, respectively, on the intestinal microcirculation which plays a crucial role in the development of multiple organ failure in sepsis. Experimental sepsis was induced in Lewis rats by intravenous lipopolysaccharide (LPS) administration. Animals were treated with simvastatin, HDL or bezafibrate. By means of intestinal intravital microscopy (IVM), the inflammatory response in the microcirculation was studied by leukocyte adherence assessment (LA) and functional capillary density (FCD) measurements. In addition, plasma levels of pro-inflammatory cytokines were determined. Bezafibrate treatment led to a reduction in leukocyte adherence, improved functional capillary density (FCD), and a reduction in interleukin-1α (IL-1α), tumour necrosis factor α (TNF-α) and granulocyte macrophage colony stimulating factors (GM-CSF) plasma levels in experimental sepsis. Contrary to this, the administration of HDL increased leukocyte adherence as well as the number of rolling leukocytes. Only IL-1α plasma levels were decreased by HDL. No significant changes were observed following simvastatin treatment. In summary, only bezafibrate showed anti-inflammatory effects in endotoxemia. This effect cannot be explained by the HDL-enhancing effect of the bezafibrate, since the direct administration of HDL showed opposite effects. Bezafibrate induced reduction of inflammation in sepsis should be investigated in further studies.

Impact of insulin on the intestinal microcirculation in a model of sepsis-related hyperglycemia.

BACKGROUND: Sepsis involves dysfunctional glucose metabolism. Among patients with sepsis, hyperglycemia is frequent and insulin administration has been evaluated for glycemic control to improve patient outcomes. Only few studies have examined the hyperglycemic microcirculation and the impact of insulin on the microvasculature in sepsis. OBJECTIVE: To study the functional capillary density (FCD) and leukocyte activation within the intestinal microcirculation in endotoxin-induced experimental sepsis. METHODS: In 50 male Lewis rats, endotoxemia was induced with lipopolysaccharide (LPS; 5 mg/kg). Low dose (LD) glucose was administered to avoid insulin-induced hypoglycemia. High dose (HD) glucose was administered to model sepsis-related hyperglycemia. Animals in LD and HD glucose groups received an insulin bolus (1.4 IU/kg). Two hours after LPS administration, intravital microscopy (IVM) of the terminal ileum was performed, and FCD and leukocyte adherence were measured in a blinded fashion. Blood glucose levels were measured every 30 min following the onset of endotoxemia. Plasma samples were collected 3 h after the onset of endotoxemia to measure IFN-γ, TNF-α, IL-1α, IL-4, GM-CSF and MCP-1 levels using multiplex bead immunoassay. RESULTS: Endotoxemia significantly reduced FCD and increased leukocyte adherence within the intestinal microvasculature. LD and HD glucose administration combined with insulin improved the FCD and decreased the adherence of leukocytes in endotoxemic animals as did HD glucose administration alone. Consistent with these results, IL-4, IL-1α, GM-CSF and IFN-γ levels were decreased following combined HD glucose and insulin administration in endotoxemic animals. CONCLUSIONS: Insulin administration, as well as an endogenous insulin response triggered by HD glucose administration, improved the FCD and decreased leukocyte activation in endotoxemic rats. The results of this study give insight into the immune and vaso-modulatory role of insulin administration during experimental endotoxemia, and may be extrapolated for clinical sepsis and other critical illnesses with marked microcirculatory dysfunction.

Experimental TLR4 inhibition improves intestinal microcirculation in endotoxemic rats.

INTRODUCTION: Toll like receptor 4 (TLR4) represents a critical cellular link for endotoxin-induced pathology. The aim of this study was to evaluate the potential role of TLR4 inhibition on the intestinal microcirculation during experimental endotoxemia. MATERIALS AND METHODS: The intestinal microcirculation was studied by intravital microscopy in four groups of Lewis rats (n=10 per group): healthy controls (CON group), endotoxemic animals (15mg/kg lipopolysaccharide, LPS group), endotoxemic animals treated with a TLR4 antagonist (1mg/kg CRX-526, LPS+CRX526 group), and controls treated with CRX-526 (C-CRX526 group). Plasma samples were obtained for cytokine measurements at the end of the experiments. RESULTS: Endotoxemia significantly increased leukocyte adhesion in intestinal submucosal venules (e.g., V1 venules: CON 20.4±6.5n/mm(2), LPS 237.5±36.2n/mm(2), p<0.05) and reduced capillary perfusion of the intestinal wall (e.g., longitudinal muscular layer: CON 112.5±5.9cm/cm(2), LPS 71.3±11.0cm/cm(2), p<0.05) at 2h. TLR4 inhibition significantly reduced endotoxemia-associated leukocyte adhesion (V1 venules: 104.3±7.8n/mm(2)) and improved capillary perfusion (longitudinal muscular layer: 111.0±12.3cm/cm(2)). Cytokine release was not significantly affected. CONCLUSIONS: The TLR4 pathway may be a target in clinical Gram-negative sepsis since administration of the TLR4 antagonist CRX-526 improved intestinal microcirculation parameters in experimental endotoxemia.

Magnesium Effects on Nonsynaptic Epileptiform Activity in Leech Retzius Neurons.

The effects of Mg2+ on Ni(2+)-induced epileptiform bursting activity and input membrane resistance during this activity of leech Retzius neurons were examined using intracellular recordings. To induce epileptiform activity, 3 mmol/l NiCl2 was added into superfusing Ringer (Ri) saline. To test for dose-dependence of the effects of Mg2+ on the induced epileptiform activity, MgCl2 was added in concentrations from 1 mmol/l to 20 mmol/l Mg2+ to the Ni(2+)-containing Ri saline. Input membrane resistance (IMR) was measured in standard Ri, Ni2+ Ri and 20 mmol/l Mg2+Ni2+ Ri saline. Superfusion with Ni2+ Ri induced epileptiform bursting activity characterized by generation of paroxysmal depolarization shifts (PDSs). Parameters of epileptiform activity including PDS frequency, PDS duration, PDS amplitude and the number of spikes/PDS were measured. Magnesium suppressed Ni(2+)-induced epileptiform activity, significantly reducing values of all parameters observed in a concentration-dependent manner. The highest concentration applied of 20 mmol/l Mg2+ completely eliminated epileptiform activity. To test for the effect of Mg2+ on membrane conductance during bursting, IMR was measured. Magnesium significantly increased IMR during bursting suppression.

Combination of dehydroepiandrosterone and orthovanadate administration reduces intestinal leukocyte recruitment in models of experimental sepsis.

BACKGROUND: Dehydroepiandrosterone (DHEA) was shown to improve the immune function and survival in experimental sepsis. This study examined the effect of DHEA on intestinal leukocyte recruitment during experimental sepsis, considering factors of gender (male, female and ovariectomized female animals) and combined treatment using orthovanadate (OV) in two models of sepsis. METHODOLOGY/FINDINGS: Male rats underwent colon ascendens stent peritonitis (CASP) or endotoxemia. DHEA was administered after induction of experimental sepsis. Changes in leukocyte adherence and capillary perfusion (measured as intestinal functional capillary density - FCD) were assessed using intravital microscopy. While DHEA increased baseline leukocyte adherence in control animals, DHEA reduced leukocyte adherence and increased FCD in male animals with CASP. These effects were also observed in DHEA-treated ovariectomized female rats with CASP. Similarly, the administration of DHEA reduced the number of adherent leukocytes to intestinal venules by 30% in the endotoxemia model. The combined treatment of DHEA and OV significantly reduced adherence of leukocytes to intestinal venules and improved FCD. CONCLUSIONS: Our results indicate that DHEA is able to reduce intestinal leukocyte recruitment induced by experimental sepsis. Combination of DHEA with OV inhibits leukocyte adherence to intestinal endothelium, similar to what is achieved by the single administration of DHEA but with significantly improved FCD. These findings suggest a potential role for DHEA and OV in clinical sepsis.

Altered basal ganglia echogenicity early in sporadic Creutzfeldt-Jakob disease.

Creutzfeldt-Jakob disease (CJD) is a fatal neurodegenerative disease caused by conformational alteration of the ubiquitous prion protein. Sporadic CJD appears to progress faster if the basal ganglia are shown to be affected on magnetic resonance imaging. Transcranial B-mode sonography (TCS) enables visualization of differences in tissue echogenicity, which can be associated with changes in the cerebral metabolism of various metals. These metabolic changes are considered 1 of the potential mechanisms of the brain damage in CJD; TCS hyperechogenicity may reflect changes in metal homeostasis in CJD. We report a 63-year-old woman who presented with typical sporadic CJD. One month after she fell ill, a magnetic resonance imaging scan of her brain showed diffuse cortical but no obvious basal ganglia involvement. However, TCS revealed moderate hyperechogenicity of both lentiform nuclei. The patient's disease progressed quickly and she died 2 months later. TCS may show basal ganglia alteration early in the disease course of patients with quickly progressing CJD, thus aiding in premortem diagnosis.

Serum Biomarker Analysis in Pediatric ADHD: Implications of Homocysteine, Vitamin B12, Vitamin D, Ferritin, and Iron Levels.

The current diagnosis of attention deficit hyperactivity disorder (ADHD) is based on history, clinical observation, and behavioral tests. There is a high demand to find biomarkers for the diagnosis of ADHD. The aim of this study is to analyze the serum profiles of several biomarkers, including homocysteine (Hcy), vitamin B12, vitamin D, ferritin, and iron, in a cohort of 133 male subjects (6.5-12.5 years), including 67 individuals with an ADHD diagnosis based on DSM-V criteria and 66 age-matched healthy boys (healthy controls, HC). Assessments for ADHD included the Iowa Conners' Teacher Rating Scale (CPRS) and the ADHDT test, as well as cognitive assessments using the Wechsler Intelligence Scale for Children-Revised (WISC-R) and the TROG-2 language comprehension test. Hcy and iron were quantified using spectrophotometry, while vitamin B12 and total 25-hydroxy vitamin D levels were determined using an electrochemiluminescence immunoassay (ECLIA) and ferritin was measured using a particle-enhanced immunoturbidimetric assay. The results showed significantly increased Hcy levels and decreased vitamin B12 levels in ADHD patients compared to HCs. Multiple logistic regression analysis indicated that Hcy is a potential prognostic indicator for ADHD. These results suggest that elevated homocysteine and decreased vitamin B12 may serve as markers for the diagnosis and prognosis of ADHD.

Estradiol receptors agonists induced effects in rat intestinal microcirculation during sepsis.

The steroid hormone estradiol is suggested to play a protective role in intestinal injury during systemic inflammation (sepsis). Our aim was to determine the effects of specific estradiol receptor (ER-α and ER-ß) agonists on the intestinal microcirculation during experimental sepsis. Male and sham ovariectomized female rats were subjected to sham colon ascendens stent peritonitis (CASP), and they were compared to male and ovariectomized female rats underwent CASP and either estradiol receptor α (ER-α) agonist propyl pyrazole triol (PPT), estradiol receptor ß (ER-ß) agonist diarylpropiolnitrile (DPN), or vehicle treatment. Intravital microscopy was performed, which is sufficiently sensitive to measure changes in the functional capillary density (FCD) as well as the major steps in leukocyte recruitment (rolling and adhesion). The leukocyte extravasations were also quantified by using histological paraffin sections of formalin fixed intestine. We found that either DPN (ER-ß) or PPT (ER-α) significantly reduced (P<0.05) sepsis-induced leukocyte-endothelial interaction (rolling, adherent leukocytes and neutrophil extravasations) and improved the intestinal muscular FCD. [PPT: Female; Leukocyte rolling (n/min): V(3) 3.7±0.7 vs 0.8±0.2, Leukocyte adhesion(n/mm(2)): V(3) 131.3±22.6 vs 57.2±13.5, Neutrophil extravasations (n/10000 µm(2)): 3.1±0.7 vs 6 ±1. Male; Leukocyte adhesion (n/mm(2)): V(1) 154.8±19.2 vs 81.3±11.2, V(3) 115.5±23.1 vs 37.8±12]. [DPN: Female; neutrophil extravasations (n/10000 µm(2)) 3.8±0.6 vs 6 ±1. Male; Leukocyte adhesion (n/mm(2)) V(1) 154.8±19.2 vs 70±10.5, V(3) 115.5±23.1 vs 52.8±9.6].Those results suggest that the observed effects of estradiol receptors on different phases of leukocytes recruitment with the improvement of the functional capillary density could partially explain the previous demonstrated salutary effects of estradiol on the intestinal microcirculation during sepsis. The observed activity of this class of compounds could open up a new avenue of research into the potential treatment of sepsis.

Activated protein C improves pial microcirculation in experimental endotoxemia in rats.

INTRODUCTION: The brain is one of the first organs affected clinically in sepsis. Microcirculatory alterations are suggested to be a critical component in the pathophysiology of sepsis. The aim of this study was to investigate the effects of recombinant human activated protein C (rhAPC) on the pial microcirculation in experimental endotoxemia using intravital microscopy. Our hypothesis is rhAPC protects pial microcirculation in endotoxemia. METHODS: Endotoxemia was generated in Lewis rats with intravenous injection of lipopolysaccharide (LPS, 5 mg/kg i.v.). Dura mater was removed through a cranial window to expose pial vessels on the brain surface. The microcirculation, including leukocyte-endothelial interaction, functional capillary density (FCD) and plasma extravasation of pial vessels was examined by fluorescent intravital microscopy (IVM) 2 h after administration of LPS, LPS and rhAPC or equivalent amount of saline (used as Control group). Plasma cytokine levels of interleukin 1 alpha (IL1-α), tumor necrosis factor-α (TNF-α), interferon γ (IFN-γ), Monocyte chemotactic protein-1 (MCP-1) and Granulocyte-macrophage colony-stimulating factor (GM-CSF) were evaluated after IVM. RESULTS: LPS challenge significantly increased leukocyte adhesion (773±190 vs. 592±152 n/mm(2) Control), decreased FCD (218±54 vs. 418±74 cm/cm(2) Control) and increased proinflammatory cytokine levels (IL-1α: 5032±1502 vs. 8±21 pg/ml; TNF-α: 1823±1007 vs. 168±228 pg/ml; IFN-γ: 785±434 vs. 0 pg/ml; GM-CSF: 54±52 vs. 1±3 pg/ml) compared to control animals. rhAPC treatment significantly reduced leukocyte adhesion (599±111 n/mm(2)), increased FCD (516±118 cm/cm(2)) and reduced IL-1α levels (2134±937 pg/ml) in the endotoxemic rats. CONCLUSION: APC treatment significantly improves pial microcirculation by reducing leukocyte adhesion and increasing FCD.

Cannabinoid receptor 2 activation reduces intestinal leukocyte recruitment and systemic inflammatory mediator release in acute experimental sepsis.

INTRODUCTION: Cannabinoid receptor 2 (CB2R) expression is upregulated during sepsis. However, there are conflicting results regarding the effects of CB2R modulation in the hyperinflammatory phase of the disease. The aim of this study was therefore to investigate the effects of CB2R manipulation on leukocyte activation within the intestinal microcirculation in two acute experimental sepsis models. METHODS: In the endotoxemia model we studied four groups of Lewis rats: controls, lipopolysaccharide (LPS), LPS + CB2R agonist HU308 (2.5 mg/kg), and LPS + CB2R antagonist AM630 (2.5 mg/kg). In the colon ascendens stent peritonitis (CASP)-induced sepsis model we also studied four groups: sham group, CASP and CASP + CB2R agonist (HU308, 2.5 or 10 mg/kg). Intravital microscopy was performed 2 hours following LPS/placebo administration or 16 hours following CASP/sham surgery to quantify intestinal leukocyte recruitment. Additionally, hemodynamic monitoring, histological examinations and measurements of inflammatory mediators were performed. RESULTS: HU308 administration significantly reduced intestinal leukocyte adhesion in both acute sepsis models. The systemic levels of inflammatory mediators were significantly reduced by 10 mg/kg HU308 treatment in CASP animals. CONCLUSION: CB2R activation reduces leukocyte activation and systemic release of inflammatory mediators in acute experimental sepsis. Drugs targeting the CB2R pathway may have therapeutic potential in sepsis.

Cannabinoid receptor 1 inhibition causes seizures during anesthesia induction in experimental sepsis.

We report on seizures during anesthesia induction in animals treated with a cannabinoid receptor 1 (CB1R) antagonist for experimental sepsis. Animals received surgery for colon ascendens stent peritonitis-induced sepsis or sham surgery followed by treatment of CB1R antagonist, CB1R agonist, or placebo. Fourteen hours later, animals received pentobarbital or ketamine for anesthesia induction and animal behavior was observed. Tonic-clonic seizures were observed in 5 of 12 septic animals (42%) treated with CB1R antagonist after induction of anesthesia with pentobarbital. The data suggest that CB1R inhibition in combination with pentobarbital may increase the incidence of anesthetic-induced seizures in the case of sepsis.

Evaluation of the effects of gender and estradiol treatment on the intestinal microcirculation during experimental sepsis.

Intestinal barrier dysfunction plays an important role in sepsis. Females may tolerate sepsis better than males, which could be due to the relative resistance of the female intestine to gut injury and inflammation when subjected to sepsis. In this study the intestinal microcirculation was investigated in 50 female and 40 male rats divided in to 9 groups of 10 animals. Male and female rats were subjected to sham CASP (colon ascendens stent peritonitis). We induced experimental sepsis (CASP) in another two groups of male and female rats. The role of the estradiol treatment was evaluated both in male and ovariectomized female rats. Female rats were subjected to sham ovariectomy 3 weeks before sham CASP. Male and ovariectomized female rats were treated with estradiol (10mg/kg estradiol in rizinus oil immediately and 12h following CASP). Another two groups of male and ovariectomized female rats received placebo oil treatment. To evaluate the effects of gender and estradiol treatment on the microvascular perfusion during sepsis, intravital microscopy was performed twenty-four hours after sham CASP or CASP surgery, which permits the in vivo determination of leukocyte-endothelial cell interaction (rolling leukocytes and adherent leukocytes) and the measurement of functional capillary density (FCD), which served as the measure of quality of microvascular perfusion. We found that there was gender difference mainly in the leukocyte endothelial interaction rather than the functional capillary density (FCD), in which male showed significant increases (P<0.05) both in the leukocyte adhesion and rolling leukocytes in submucosal venules (V1 and V3) in comparison to female rats. (Leukocyte adhesion: V1 107.1 ± 49.2n/mm(2); V3 112.3 ± 68.1n/mm(2)) (Rolling leukocytes:V1 16.2 ± 10.3n/min; V3 8.4 ± 8.2n/min). In addition estradiol replacement in ovariectomized female and male rats induced significant decreases (P<0.05) in the leukocyte adhesion and rolling (V1 and V3) with a significant increase in the muscular FCD in comparison to the corresponding placebo treated groups. (Leukocyte adhesion: V1 60 ± 31 n/mm(2); V3 78.11 ± 37.6n/mm(2)) (Rolling leukocytes: V1 13.4 ± 8.9 n/min; V3 5.8 ± 7.4n/min) (Longitudinal muscular FCD (cm/cm(2)): in ovariectomized female rats 107.1 ± 12.2; in male rats 106.2 ± 15.3) (Circular muscular FCD (cm/cm(2)): in ovariectomized female rats: 84.8 ± 14.2; in male rats: 86.1 ± 15.3). We conclude that gender difference in the leukocyte endothelial interaction could explain the resistance of female intestine to injury, and that estradiol treatment could improve the intestinal microcirculation through its effects on the leukocyte endothelial interaction and the FCD both in male and ovariectomized female rats.

The expression of myogenic regulatory factors and muscle growth factors in the masticatory muscles of dystrophin-deficient (mdx) mice.

The activities of myogenic regulatory factors (MRF) and muscle growth factors increase in muscle that is undergoing regeneration, and may correspond to some specific changes. Little is known about the role of MRFs in masticatory muscles in mdx mice (the model of Duchenne muscular dystrophy) and particularly about their mRNA expression during the process of muscle regeneration. Using Taqman RT-PCR, we examined the mRNA expression of the MRFs myogenin and MyoD1 (myogenic differentiation 1), and of the muscle growth factors myostatin, IGF1 (insulin-like growth factor) and MGF (mechano-growth factor) in the masseter, temporal and tongue masticatory muscles of mdx mice (n = 6 to 10 per group). The myogenin mRNA expression in the mdx masseter and temporal muscle was found to have increased (P < 0.05), whereas the myostatin mRNA expressions in the mdx masseter (P < 0.005) and tongue (P < 0.05) were found to have diminished compared to those for the controls. The IGF and MGF mRNA amounts in the mdx mice remained unchanged. Inside the mdx animal group, gender-related differences in the mRNA expressions were also found. A higher mRNA expression of myogenin and MyoD1 in the mdx massterer and temporal muscles was found in females in comparison to males, and the level of myostatin was higher in the masseter and tongue muscle (P < 0.001 for all comparisons). Similar gender-related differences were also found within the control groups. This study reveals the intermuscular differences in the mRNA expression pattern of myogenin and myostatin in mdx mice. The existence of these differences implies that dystrophinopathy affects the skeletal muscles differentially. The finding of gender-related differences in the mRNA expression of the examined factors may indicate the importance of hormonal influences on muscle regeneration.

Differential expression of myosin heavy chain isoforms in the masticatory muscles of dystrophin-deficient mice.

The dystrophin-deficient mouse (mdx) is a homologue animal model of Duchenne muscular dystrophy (DMD) and is characterized by slowly progressive muscle weakness accompanied by changes in myosin heavy chain (MyHC) composition. It is likely that the masticatory muscles undergo similar changes. The aim of this study was to examine the masticatory muscles (masseter, temporal, tongue, and soleus) of 100-day-old mdx and control mice (n = 8-10), and the fibre type distribution (by immunohistochemistry) as well as the expression of the corresponding MyHC messenger RNA (mRNA) (protein and mRNA expression, using Western blot or quantitative real-time polymerase chain reaction (RT-PCR)). Immunohistochemistry and western blot analysis revealed that the masticatory muscles in the control and mdx mice consisted mainly of type 2 fibres, whereas soleus muscle consisted of both type 1 and 2 fibres. In the masseter muscle, the mRNA in mdx mice was not different from that found in the controls. However, the mRNA content of the MyHC-2b isoform in mdx mice was lower in comparison with the controls in the temporal muscle [11.9 versus 36.9 per cent; P < 0.01; mean ± standard error of the mean (SEM), Student's unpaired t-test], as well as in the tongue muscle (65.7 versus 73.8 per cent; P < 0.05). Similarly, the content of MyHC-2x isoforms in mdx tongue muscle was lower than in the controls (25.9 versus 30.8 per cent; P < 0.05). The observed down-regulation of the MyHC-2x and MyHC-2b mRNA in the masticatory muscles of mdx mice may lead to changed fibre type composition. The different MyHC gene expression in mdx mice masticatory muscles may be seen as an adaptive mechanism to muscular dystrophy.