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Spinal muscular atrophy (SMA) is a genetically inherited recessive neuromuscular disease that causes muscular atrophy and weakness. Onasemnogene abeparvovec (formerly AVXS-101, Zolgensma®, Novartis) is a targeted therapy approved to treat patients with SMA in >40 countries worldwide. This study describes the clinical efficacy and tolerability of gene replacement therapy with onasemnogene abeparvovec over a 3-month period in 9 SMA type 1 patients aged 1.7-48 months, with 7 patients on stable nusinersen (i.e., had received all four nusinersen loading doses before inclusion in this study). Liver function (alanine aminotransferase, aspartate aminotransferase, total bilirubin), troponin I, platelet counts, creatinine levels, and motor function (CHOP-INTEND) were monitored. For the seven patients on stable nusinersen, the median baseline CHOP-INTEND score increased significantly during nusinersen treatment (Wilcoxon signed-rank test p = 0.018) and at 3 months after switching to onasemnogene abeparvovec (Wilcoxon signed-rank test p = 0.0467). We also identified two patients who responded poorly to nusinersen but showed the largest increase in baseline CHOP-INTEND scores at 1 and 3 months after switching, which could suggest that poor responders to nusinersen may respond favorably to onasemnogene abeparvovec. No unknown adverse events occurred. One patient developed moderate/severe thrombocytopenia 1 week after onasemnogene abeparvovec administration that resolved after treatment. Our study suggests the possibility of a change in the dynamic of CHOP-INTEND for patients who respond poorly to nusinersen after switching therapy to onasemnogene abeparvovec. Alternatively, patient age at treatment initiation may impact the response to onasemnogene abeparvovec. Testing in larger patient populations must be undertaken to assess the plausibility of these hypotheses., (© 2022. The Author(s).)

The epidemic spread of childhood obesity in Western society has interested many researchers, who agree in defining it as a multifactorial disease in which not only eating habits and sedentary lifestyle play a role, but also genetic predisposition. The aim of this study was to analyze the personality profile of a group of mothers of children with obesity and to compare this profile to that of a group of mothers of children without obesity. A total of 258 mothers participated in the study (126 mothers of children with obesity and 132 mothers of children without obesity). Weight and height were measured and the body mass index was calculated. The Minnesota Multiphasic Personality Inventory second edition (MMPI-2), evaluating personality and psychological disorders, was used to evaluate the personality profile. The results suggested that mothers of children with obesity score higher than the mothers of children without obesity in all MMPI-2 subscales. In most of these subscales, the differences between the two groups of mothers were statistically significant and with a medium to high effect size. These data suggest a new perspective on childhood obesity, identifying it as a multifactorial pathology that requires a multimodal and multidisciplinary approach that also takes care of caregivers to ensure optimal therapeutic efficacy.

Introduction: In eating disorders, the association with other psychiatric symptoms is of particular interest. The association between anorexia nervosa and psychotic symptoms is less studied than that with affective disorders (anxiety/depression). The aim of this study is to describe a psychotic symptom (paranoia) in adolescents with anorexia nervosa looking at several potential explicative associated factors: eating disorder symptoms, body image concerns, depression and social anxiety. Our hypothesis is that paranoia in anorexia nervosa patients is better explained by the concomitant depression and social anxiety symptoms than the core symptoms of the disease (eating disorder symptoms or body image concerns)., Methods: This is a retrospective study. Consecutive, help-seeking adolescents, admitted to the Eating Disorder Service of the Integrated Pediatric Care Department, Luigi Vanvitelli University Hospital, constituted the sample. Data was obtained through retrospective collection of clinical interviews and self-report questionnaires, used for the routine assessment of these patients, administered by trained and expert child and adolescent psychiatrists, they were the Eating Attitude Test-26 (EAT-26), the Body Uneasiness Test-A (BUT-A), the Children Depression Inventory (CDI), the Liebowitz Social Anxiety Scale-Children and Adolescents (LSAS-CA) and the paranoia subscale of the Specific Psychotic Experiences Questionnaire (SPEQ)., Results: We obtained data from 92 adolescents with anorexia nervosa and other specified feeding or eating disorder (OSFED). Our regression model explained that paranoia (SPEQ-paranoia subscale) in this population was better explained by depression (CDI) (coefficient= 0.415 SD: 0.210, p=0.052) and social anxiety symptoms (LSAS-CA) (coefficient= 0.253 SD: 0.060; p<0.001) than eating disorder symptoms (EAT-26) (coefficient= 0.092 SD: 0.107; p=0.398) and body image concerns (BUT-A) (coefficient= 1.916 SD: 2.079; p=0.359)., Conclusions: This study has some theoretical, clinical and treatment implications. It is important to carrying out screening for the presence of psychotic symptoms in patients with eating or feeding disorders. These symptoms and associated factors (depression and social anxiety) may complicate the clinical picture of the disease with the need, in certain cases, of psychopharmacological drugs and, among these, anti-psychotics. Finally in the psychotherapy context, paranoid ideas may be subject of treatment for patient with anorexia nervosa.

Neurodevelopmental disorders (NDDs) include diverse neuropathologies characterized by abnormal brain development leading to impaired cognition, communication and social skills. A common feature of NDDs is defective synaptic plasticity, but the underlying molecular mechanisms are only partially known. Several studies have indicated that people's lifestyles such as diet pattern and physical exercise have significant influence on synaptic plasticity of the brain. Indeed, it has been reported that a high-fat diet (HFD, with 30-50% fat content), which leads to systemic low-grade inflammation, has also a detrimental effect on synaptic efficiency. Interestingly, metabolic alterations associated with obesity in pregnant woman may represent a risk factor for NDDs in the offspring. In this review, we have discussed the potential molecular mechanisms linking the HFD-induced metabolic dysfunctions to altered synaptic plasticity underlying NDDs, with a special emphasis on the roles played by synaptic protein synthesis and mitochondrial functions.

Background and Objectives: Siblings of disabled children are more at risk of developing mental illnesses. More than 50 international studies show that about 8% of children and adolescents suffer from a mental disorder, which is almost always a source of difficulties both at the interpersonal level (in the family and with peers) and at school. Healthy siblings of children with disabilities are one of the groups most at risk for consequences in psychological health and well-being. As some authors suggest, siblings build their idea of "being people", in terms of character and personality, by continuously and daily confronting themselves with the theme of disability and a family context subjected to continuous stress. The following contribution aims to compare emotional-behavioral disorders in healthy siblings of children with autism spectrum disorder, in healthy siblings of children with Down's syndrome and in healthy siblings of children with typical development. Materials and Methods : The results involve 153 children from the region of Campania and their caregivers through the administration of the Strength and Difficulties Questionnaire. Results : From the data, it emerged that siblings of children with autism spectrum disorder and siblings of children with Down's syndrome have a greater emotional fragility, especially among male subjects. Conclusions: Our results require us to reflect on the clinical and policy measures needed to ensure the well-being of siblings of disabled children, mainly through appropriate sibling coping training.

A large body of literature reports the higher prevalence of epilepsy in subjects with Autism Spectrum Disorder (ASD) compared to the general population. Similarly, several studies report an increased rate of Subclinical Electroencephalographic Abnormalities (SEAs) in seizure-free patients with ASD rather than healthy controls, although with varying percentages. SEAs include both several epileptiform discharges and different non-epileptiform electroencephalographic abnormalities. They are more frequently associated with lower intellectual functioning, more serious dysfunctional behaviors, and they are often sign of severer forms of autism. However, SEAs clinical implications remain controversial, and they could represent an epiphenomenon of the neurochemical alterations of autism etiology. This paper provides an overview of the major research findings with two main purposes: to better delineate the state-of-the-art about EEG abnormalities in ASD and to find evidence for or against appropriateness of SEAs pharmacological treatment in ASD.

Functional non-retentive fecal incontinence (FNRFI) is a common problem in pediatric age. FNRFI is defined as unintended loss of stool in a 4-year-old or older child after organic causes have been excluded. FNRFI tends to affects up to 3% of children older than 4 years, with males being affected more frequently than females. Clinically, children affected by FNRFI have normal intestinal movements and stool consistency. Literature data show that children with fecal incontinence have increased levels of separation anxiety, specific phobias, general anxiety, attention-deficit/hyperactivity disorder (ADHD), and oppositional defiant disorder. In terms of possible relationship between incontinence and sleep, disorders of sleep organization have been observed in the pathogenesis of enuresis so generating the hypothesis that the orexinergic system may have a crucial role not only for the sleep organization per se but also for the sphincterial control in general. This study aimed to focus on specific neurophysiological aspects to investigate on the possible relationship between sleep organizational abnormalities and FNRFI. Specifically, we aimed to measure orexin serum levels in children with FNRFI and assess their polysomnographic sleep macrostructure patterns. Two study groups were considered: FNFRI ( n = 45) and typically developed (TD) ( n = 45) group. In both groups, sleep patterns and respiratory events were assessed by polysomnographic recordings (PSG) during a period of two nights at least, and plasma levels of Orexin-A were measured in each participant. The findings of this initial investigation seem to support a major role of Orexin-A in sleep organization alterations in children with FNFRI. Also, our data suggest that sleep habits evaluation should be considered as screening and complementary tool for the diagnosis of fecal incontinence in children.

Background: Primary monosymptomatic nocturnal enuresis (PMNE) may have a stressful impact on the everyday life of children and parents, and it may represent a cumulative stress factor increasing feelings of "learned helplessness." Methods: The current study investigated parental stress in a group of parents ( n = 330) of children affected by PMNE, compared to a group of parents ( n = 330) of typical developing children (TDC). In addition, the study evaluated whether parents of PMNE children experience more emotional, social, and behavioral problems in their children, compared to parents of TDC. Finally, the study correlated frequency of enuresis with stress values and Child Behavior Checklist (CBCL) subscales and total stress with CBCL. Both groups were given The Parental Stress Inventory-Short Form (PSI-SF) and the Child Behavior Checklist (CBCL). Results: Parents of PMNE children showed significantly higher stress level than parents of TDC. Nocturnal enuresis, as a demanding clinical condition difficult to control, represents a relevant stress factor. Mothers appeared as more vulnerable to stress than fathers. Parents of PMNE children reported higher behavioral and emotional problems, compared to reports of parents of TDC. PMNE children appeared to their parents as having lower competency in social activities, school performance, and social relationships than TDC. Moreover, they were rated as more withdrawn, anxious-depressed, more aggressive, inattentive, and with more somatic complaints than healthy children. It was always the mother who rated a significantly higher number of emotional, social, and behavioral problems compared to fathers. Correlational analysis showed that the higher the frequency of enuresis, the greater the parental stress level, the lower the social activities, school performance and relational competencies and the higher the emotional, social and behavioral problems in children, according to the parents' evaluations. The greater the parental stress level, the lower the competencies rated and the higher the behavioral problems detected by parents. Conclusion: The physicians who deal with PMNE children have taken into account the stressful role and emotional dimensions of this clinical condition, both for children and mothers, in order to improve clinical management. Psychological support is needed for parents, and mothers especially, for a more functional stress management related to the PMNE., (Copyright © 2019 Roccella, Smirni, Smirni, Precenzano, Operto, Lanzara, Quatrosi and Carotenuto.)

Developmental Coordination Disorder (DCD) is considered to be abnormal motor skills learning, identified by clumsiness, slowness, and/or motor inaccuracy impairing the daily-life activities in all ages of life, in the absence of sensory, cognitive, or neurological deficits impairment. The present research focuses on studying DCD sleep structure and Cyclic Alternating Pattern (CAP) parameters with a full overnight polysomnography and to study the putative correlations between sleep architecture and CAP parameters with motor coordination skills. The study was a cross-sectional design involving 42 children (26M/16F; mean age 10.12 ± 1.98) selected as a DCD group compared with 79 children (49M/30F; mean age 9.94 ± 2.84) identified as typical (no-DCD) for motor ability and sleep macrostructural parameters according to the MABC-2 and polysomnographic (PSG) evaluations. The two groups (DCD and non-DCD) were similar for age ( p = 0.715) and gender ( p = 0.854). More significant differences in sleep architecture and CAP parameters were found between two groups and significant correlations were identified between sleep parameters and motor coordination skills in the study population. In conclusion, our data show relevant abnormalities in sleep structure of DCD children and suggest a role for rapid components of A phases on motor coordination development.

Background: Obstructive sleep apnea syndrome (OSAS) affects up to 4% of a pediatric population, with many comorbidities in the medium-long term. Functional alterations in the prefrontal cortex (PFC) may explain why OSAS impacts aspects such as executive functions, memory, motor control, attention, visual-spatial skills, learning, and mood regulation. Emotional intelligence (EI) is a complex neuropsychological function that could be impaired in many clinical conditions., Purpose: The aim of the study is to evaluate the difference in emotional intelligence skills among children with OSAS and healthy subjects (nOSAS)., Methods: 129 children (72 males; mean age 7.64 ± 1.98 years) affected by OSAS were compared to 264 non-OSAS (nOSAS) children (138 males; mean age 7.98 ± 2.13) similar for gender, age, and socioeconomic status. In order to assess the emotional quotient, the Bar-On Emotional Quotient Inventory: Youth Version (EQ-i:YV) was used., Results: The comparison for means and standard deviation between OSAS children and nOSAS children for EQ-i:YV scores showed significant differences for Interpersonal, Adaptability, and Stress Management scales and EQ Total score., Conclusions: Our findings highlighted the role of intermittent hypoxia in the genesis of the effects of sleep-related respiratory disorders, which involves also aspects different from physical impairments., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2019 Francesca Felicia Operto et al.)

Background: Studies about the impact of developmental dyslexia (DD) on parenting are scarce. Our investigation aimed to assess maternal stress levels and mothers' copying styles in a population of dyslexic children., Methods: A total of 874 children (500 boys, 374 girls; mean age 8.32 ± 2.33 years) affected by DD was included in the study. A total of 1,421 typically developing children (789 boys, 632 girls; mean age 8.25 ± 3.19 years) were recruited from local schools of participating Italian Regions (Abruzzo, Calabria, Campania, Puglia, Umbria, Sicily) and used as control-children group. All mothers (of both DD and typically developing children) filled out an evaluation for parental stress (Parenting Stress Index-Short Form) and coping strategies [Coping Inventory for Stressful Situations (CISS)]., Results: No statistical differences for mean age ( p  = 0.456) and gender ( p  = 0.577) were found between DD and control children. Mothers of children affected by DD showed an higher rate of all parental stress indexes (Parental Distress domain p  < 0.001, Difficult Child p  < 0.001, Parent-Child Dysfunctional Interaction p  < 0.001, and Total Stress subscale score p  < 0.001) than controls mothers. According to the CISS evaluation, mothers of DD children reported a significantly higher rate of emotion-oriented ( p  < 0.001) and avoidance-oriented ( p  < 0.001) coping styles than mothers of typical developing children. On the other hand, a lower representation of task-oriented coping style was found in mothers of DD children ( p  < 0.001) in comparison to mothers of control-children., Conclusion: Our study shows the clinical relevance of the burden carried by the mothers of children affected by DD and suggests the importance to assess parents, particularly mothers, to improve family compliance and clinical management of this disorder.

During the past decade, a substantial increase in the use of second generation antipsychotics (SGAs) has occurred for a number of juvenile psychiatric disorders, often as off-label prescriptions. Although they were thought to be safer than older, first generation antipsychotics, mainly due to a lower risk of neurological adverse reactions, recent studies have raised significant concerns regarding their safety regarding metabolic, endocrinological and cardiovascular side effects. Aim of this paper is to update with a narrative review, the latest findings on safety of SGAs in youths. Results suggest that different SGAs may present different safety profiles. Metabolic adverse events are the most frequent and troublesome, with increasing evidences of heightened risk for type II diabetes mellitus. Results are discussed with specific emphasis on possible strategies of an active monitoring, which could enable both paediatricians and child psychiatrists to a possible prevention, early detection, and a timely management of such effects.

The aim of the present study was to analyze the protein composition of ductal breast carcinoma and the surrounding normal tissue in individual patients using comparative 2D proteomics and mass spectrometry to detect candidate disease biomarkers for diagnosis and prognosis. Samples of normal and cancerous tissue obtained form 28 patients were analyzed. Chaperonins and cytoskeletal proteins predominated among the 11 proteins for which major changes in abundance were detected. Of these 11 proteins with an altered expression, 2 had a decreased expression and 9 had an increased expression. In addition, the abundance of a few cytokeratins was also altered; however, they were not capable of serving as specific circulatory biomarkers. The proteins which we observed to exhibit an altered expression in infiltrating ductal breast carcinoma may be exploited as novel targets for therapeutic interventions or represent novel diagnostic/prognostic markers for the early detection of aggressive tumors, particularly those with multridrug-resistant phenotypes during the earlier stages of the disease.

Tissue transglutaminase undergoes thermal inactivation with first-order kinetics at moderate temperatures, in a process which is affected in opposite way by the regulatory ligands calcium and GTP, which stabilize different conformations. We have explored the processes of inactivation and of unfolding of transglutaminase and the effects of ligands thereon, combining approaches of differential scanning calorimetry (DSC) and of thermal analysis coupled to fluorescence spectroscopy and small angle scattering. At low temperature (38-45°C), calcium promotes and GTP protects from inactivation, which occurs without detectable disruption of the protein structure but only local perturbations at the active site. Only at higher temperatures (52-56°C), the protein structure undergoes major rearrangements with alterations in the interactions between the N- and C-terminal domain pairs. Experiments by DSC and fluorescence spectroscopy clearly indicate reinforced and weakened interactions of the domains in the presence of GTP and of calcium, and different patterns of unfolding. Small angle scattering experiments confirm different pathways of unfolding, with attainment of limiting values of gyration radius of 52, 60 and 90 Å in the absence of ligands and in the presence of GTP and calcium. Data by X-rays scattering indicate that ligands influence retention of a relatively compact structure in the protein even after denaturation at 70°C. These results suggest that the complex regulation of the enzyme by ligands involves both short- and long-range effects which might be relevant for understanding the turnover of the protein in vivo.

The transamidating activity of tissue transglutaminase is regulated by the ligands calcium and GTP, via conformational changes which facilitate or interfere with interaction with the peptidyl-glutamine substrate. We have analysed binding of these ligands by calorimetric and computational approaches. In the case of GTP we have detected a single high affinity site (K (D) approximately 1 microM), with moderate thermal effects suggestive that binding GTP involves replacement of GDP, normally bound to the protein. On line with this possibility no significant binding was observed during titration with GDP and computational studies support this view. Titration with calcium at a high cation molar excess yielded a complex binding isotherm with a number of "apparent binding sites" in large excess over those detectable by equilibrium dialysis (6 sites). This binding pattern is ascribed to occurrence of additional thermal contributions, beyond those of binding, due to the occurrence of conformational changes and to catalysis itself (with protein self-crosslinking). In contrast only one site for binding calcium with high affinity (K (D) approximately 0.15 microM) is observed with samples of enzyme inactivated by alkylation at the active site (to prevent enzyme crosslinkage and thermal effects of catalysis). These results indicate an intrinsic ability of tissue transglutaminase to bind calcium with high affinity and the necessity of careful reassessment of the enzyme regulatory pattern in relation to the concentrations of ligands in living cells, taking also in account effects of ligands on protein subcellular compartimentation.

Activation of tissue transglutaminase by calcium involves a conformational change which allows exposition of the active site to the substrate via movements of domains 3 and 4 that lead to an increase of the inter-domain distance. The inhibitor GTP counteracts these changes. Here we investigate the possible existence of non-native conformational states still compatible with the enzyme activity produced by chemical and thermal perturbations. The results indicate that chemical denaturation is reversible at low guanidine concentrations but irreversible at high concentrations of guanidine. Indeed, at low guanidine concentrations tissue TG-ase exists in a non-native state which is still affected by the ligands as in the native form. In contrast, thermal unfolding is always irreversible, with aggregation and protein self-crosslinkage in the presence of calcium. DSC thermograms of the native protein in the absence of ligands consist of two partly overlapped transitions, which weaken in the presence of calcium and merge together and strengthen in the presence of GTP. Overall, the present work shows, for the first time, the reversible denaturation of a TG-ase isoenzyme and suggests the possibility that also in in vivo, the enzyme may acquire non-native conformations relevant to its patho-physiological functions.

Background: Severe chronic venous disease (CVD) is characterized by both dermal hemosiderin accumulation and matrix metalloproteinase (MMP) hyperactivation. The iron-driven pathway is one of the recognized mechanisms of MMP hyperactivation., Objective: To investigate the potential consequences of leg hemosiderin deposits on both iron metabolism and activation of MMPs., Methods: We contemporaneously assessed the following in the serum of the arm and ankle veins of 30 patients (C4-6) with CVD and 14 normal subjects: ferritin, transferrin, iron, percentage of transferrin iron binding capacity (%TIBC), and MMP-9. Optical microscopy examinations with Perls' staining of chronic wounds were also performed., Results: Histology consistently revealed iron deposits. Serum ferritin, iron, and %TIBC were significantly increased in the legs affected by severe CVD compared with the arm of the same subjects or the controls. In addition, iron and %TIBC were significantly elevated in the legs of ulcer patients. The rate of activation of MMP-9 was significantly elevated in CVD., Conclusions: The increased iron deposition in legs affected by CVD seems to be more instable in ulcer patients, leading to iron release in the serum of the affected leg. Our data suggest the iron-driven pathway as a further mechanism for MMP hyperexpression leading to tissue lesion.

Objective: Impaired venous drainage in severe chronic venous insufficiency (CVI) leads to microcirculatory overload, characterized by erythrocyte diapedesis and subsequent extravascular hemolysis, resulting in typical dermal hemosiderin deposition. We hypothesized that hemosiderin, normally absent, could be present in the urine in CVI., Methods: The three-phase study included 117 patients with CVI and 12 healthy control subjects, all of whom had undergone clinical examination and duplex scanning. In phase 1, current methods were used to test urine for hemosiderin in 61 persons: 12 healthy control subjects, 24 patients with mild CVI (clinical class C1 to C3), and 25 patients with severe CVI (clinical class C4 to C6). In phase 2, the concentration of urinary hemosiderin was determined in 45 consecutive patients with CVI, CEAP class 1 to 6. A score of 0 was assigned when typical hemosiderin granules were absent at microscopic examination, a score of 1 when one to three granules per field were detected; 2 when four to six granules were detected; and 3 when more than six granules were observed. Phase 3 included 23 patients with CVI (clinical class 2 to 6). Hemosiderin concentration was determined and a score assigned before patients underwent surgical procedures to correct primary CVI. Both hemosiderin testing and duplex scanning were repeated after 6 months., Results: Phase 1: Urine hemosiderin testing to determine presence or absence of CVI in patients with reflux detectable at duplex scanning yielded the following values: positive predictive value, 96% (95% confidence interval [CI], 86% to 100%); negative predictive value, 88% (CI, 68% to 97%); sensitivity, 94% (CI, 72% to 99%); specificity, 91% (CI, 83% to 99%); and diagnostic accuracy, 95% (CI, 86% to 99%). Phase 2: Hemosiderinuria score enabled classification of clinical severity of CVI. Mean scores, respectively, were clinical class 1, 0.18 +/- 0.12; class 2, 0.75 +/- 0.47; class 3, 1.67 +/- 0.21; class 4, 1.86 +/- 0.26; class 5, 2.50 +/- 0.28; and class 6, 1.92 +/- 0.21 (P <.001). Phase 3: At 6-month follow-up, hemosiderin score was improved, from 2.48 +/- 0.12 preoperatively to 0.78 +/- 0.18 postoperatively (P <.0001). A score of 0 or 1 was associated with successful surgery, whereas a score of 2 or 3 reflected persistence of reflux., Conclusions: Determination of presence of hemosiderin in the urine is a new, sensitive, cost-effective, noninvasive, and repeatable test that enables detection of substantial microcirculatory overload in patients with CVI.

Purified bile salt hydrolase from bile-adapted Xanthomonas maltophilia displays Michaelis-Menten kinetics on cholylglycine and cholyltaurine and hydrolyzes bile salts also in crude bovine bile. The protein is a dimer and is resistant to proteinases and to heating at 55 to 60 degrees C for up to 60 min, in agreement with calorimetric data.

Tissue-type transglutaminase is inactivated in a time-dependent way during incubation with submillimolar concentrations of o-phthalaldehyde, with affinity labeling kinetics. The rate of inactivation by the reagent is greatly enhanced in the presence of the essential enzyme cofactor calcium and is decreased by GTP, an allosteric inhibitor. A fluorescent isoindole derivative is formed during the modification apparently through crosslinkage of active site Cys 277 to a lysine residue. These data and the quenching of fluorescence by addition of calcium ions suggest that the enzyme active site is directly involved in the inactivation process.

Homogenates of Yoshida hepatoma cells, cultured as ascite suspension in vivo, display significant transglutaminase activity in both the cytosolic and the particulate fraction. The enzyme, however, is predominantly membrane-bound. Transglutaminase was solubilized from the membranes either by extraction with detergents or treatment with neutralized hydroxylamine or proteinases. We observed similar molecular weight under denaturing conditions, catalytic and immunologic properties for purified cytosolic and solubilized transglutaminase, and identity of the limited proteolytic maps. These results suggest that transglutaminase isoforms actually consist of the same protein undergoing translocation by unknown mechanisms.

A simple technique is described for increasing the number of mitoses in circulating blood and in the kidney of teleost fish. This method employed phytohaemagglutinin-P solution, a T-like cell mitogen, which was injected (1 mg/100 g body wt) intraperitoneally, and 53 h later many blasts suitable for karyological studies, blocked in metaphase, were present both in the circulating blood and in the kidney.

We have studied the effect of sonication on the fluorescence of N-(1-pyrenyl)iodoacetamide-labeled F-actin as well as of native actin-pyrenyl-actin mixed oligomers in which the subunits were covalently attached to each other by phenylenebismaleimide. In both cases the fluorescence of the solution was largely decreased by sonication. We have found that this effect is due (a) to a 20-30% decrease of the specific fluorescence of the polymers. These results question the validity of the novel mechanism for the polymerization of actin recently proposed (D. Pantaloni et al. (1984) J. Biol. Chem. 259, 6274-6283). In these studies, in fact, the implicit assumption was made that the quenching of the fluorescence of the solution under sonication was due exclusively to the conversion of F-actin into G-actin.

At or below -12 degrees C and in the presence of 40% ethylene glycol, only two out of the four dihydroxyacetone phosphate binding sites of aldolase are catalytically active. At these same temperatures and at pH* 8.3, the equilibrium between the pre-enamine and the enamine plus the post-enamine intermediates is largely shifted in favor of the latter. The enamine phosphate and the enamine-aldehyde phosphate intermediates have been resolved by studying the rate of their formation at -13 degrees C and pH* 5.28 and the trapping by DL-glyceraldehyde 3-phosphate at -24 degrees C and pH* 5.24.

In the presence of Mg2+, the formation of actin filaments is hindered by glyceraldehyde-3-phosphate dehydrogenase. This effect, which increases with the square of Mg2+ concentration, is counteracted by 0.15 M KCl. Thus KCl, at concentrations found in the intracellular compartment, appears to be strictly required for the correct formation of actin filaments in all tissues in which the glyceraldehyde-phosphate dehydrogenase concentration is high.

Xanthine oxidase increases the rate of actin polymerization. This occurs at oxidase concentrations as low as 40 nM provided the concentration of the polymerizing agent is low (0.5 mM MgCl2). In the presence of 0.1 M KCl plus 1 mM MgCl2 as the polymerizing agents, xanthine oxidase does not affect the rate of the polymerization but increases significantly the rate of the conversion of F(ATP)actin into F(ADP.Pi)actin and probably also the rate of the orthophosphate release.