Variant-specific effects of GBA1 mutations on dopaminergic neuron proteostasis.

Glucocerebrosidase 1 (GBA1) mutations are the most important genetic risk factors for Parkinson's disease (PD). Clinically, mild (e.g., p.N370S) and severe (e.g., p.L444P and p.D409H) GBA1 mutations have different PD phenotypes, with differences in age at disease onset, progression, and the severity of motor and non-motor symptoms. We hypothesize that GBA1 mutations cause the accumulation of α-synuclein by affecting the cross-talk between cellular protein degradation mechanisms, leading to neurodegeneration. Accordingly, we tested whether mild and severe GBA1 mutations differentially affect the degradation of α-synuclein via the ubiquitin-proteasome system (UPS), chaperone-mediated autophagy (CMA), and macroautophagy and differentially cause accumulation and/or release of α-synuclein. Our results demonstrate that endoplasmic reticulum (ER) stress and total ubiquitination rates were significantly increased in cells with severe GBA1 mutations. CMA was found to be defective in induced pluripotent stem cell (iPSC)-derived dopaminergic neurons with mild GBA1 mutations, but not in those with severe GBA1 mutations. When examining macroautophagy, we observed reduced formation of autophagosomes in cells with the N370S and D409H GBA1 mutations and impairments in autophagosome-lysosome fusion in cells with the L444P GBA1 mutation. Accordingly, severe GBA1 mutations were found to trigger the accumulation and release of oligomeric α-synuclein in iPSC-derived dopaminergic neurons, primarily as a result of increased ER stress and defective macroautophagy, while mild GBA1 mutations affected CMA, which is mainly responsible for the degradation of the monomeric form of α-synuclein. Overall, our findings provide new insight into the molecular basis of the clinical variability in PD associated with different GBA1 mutations.

A matter of the heart: myocardial metastases in neuroendocrine tumors.

The aim of the present study was to evaluate frequency, clinical spectrum, and treatment of myocardial metastases in patients with histologically proven neuroendocrine tumors by analysis of our database and literature review. The literature on cardiac metastases in patients with neuroendocrine tumors published from 1973 to the present was reviewed for age, sex, primary tumor localization, metastases, symptoms, complications, treatment, diagnostic methods, and histology. Patient records from our institution were analyzed retrospectively for cardiac metastases detected by any diagnostic means and detailed patient histories are given. 4 patients with myocardial metastases could be identified in our database (n=550) while literature review identified 41 published cases. Mean age at initial diagnosis was 57.5 years (females=13, males=28), primary tumor localizations were foregut (n=7), midgut (n=28), hindgut (n=1), or unknown (n=3). Carcinoid syndrome was reported for 28 patients. Cardiac involvement was right-ventricular only (n=10), left-ventricular only (n=11), or biventricular (n=10). Diagnosis was obtained by echocardiography (n=21), CT/MRI (n=12) and other methods (n=9), or by autopsy (n=9). We describe visualization of cardiac metastases by (68)Ga-DOTATOC-PET/CT for the first time. Clinical presentation ranged from asymptomatic patients to cardiac arrest. Follow-up times ranged from <1 month up to 12 years. Clinicians treating patients with neuroendocrine tumors should be aware of the heart as a possible site of metastatic disease. Echocardiography and MRI are the methods of choice for follow-up, while PET/CT might contribute to earlier and more frequent detection. Management of cardiac metastases requires close cooperation between specialists of internal medicine, nuclear medicine, and cardiac surgery.

Cardiomyopathy mutations reveal variable region of myosin converter as major element of cross-bridge compliance.

The ability of myosin to generate motile forces is based on elastic distortion of a structural element of the actomyosin complex (cross-bridge) that allows strain to develop before filament sliding. Addressing the question, which part of the actomyosin complex experiences main elastic distortion, we suggested previously that the converter domain might be the most compliant region of the myosin head domain. Here we test this proposal by studying functional effects of naturally occurring missense mutations in the beta-myosin heavy chain, 723Arg --> Gly (R723G) and 736Ile --> Thr (I736T), in comparison to 719Arg --> Trp (R719W). All three mutations are associated with hypertrophic cardiomyopathy and are located in the converter region of the myosin head domain. We determined several mechanical parameters of single skinned slow fibers isolated from Musculus soleus biopsies of hypertrophic cardiomyopathy patients and healthy controls. Major findings of this study for mutation R723G were i), a >40% increase in fiber stiffness in rigor with a 2.9-fold increase in stiffness per myosin head (S( *)(rigor R723G) = 0.84 pN/nm S( *)(rigor WT) = 0.29 pN/nm); and ii), a significant increase in force per head (F( *)(10 degrees C), 1.99 pN vs. 1.49 pN = 1.3-fold increase; F( *)(20 degrees C), 2.56 pN vs. 1.92 pN = 1.3-fold increase) as well as stiffness per head during isometric steady-state contraction (S( *)(active10 degrees C), 0.52 pN/nm vs. 0.28 pN/nm = 1.9-fold increase). Similar changes were found for mutation R719W (2.6-fold increase in S( *)(rigor); 1.8-fold increase in F( *)(10 degrees C), 1.6-fold in F( *)(20 degrees C); twofold increase in S( *)(active10 degrees C)). Changes in active cross-bridge cycling kinetics could not account for the increase in force and active stiffness. For the above estimates the previously determined fraction of mutated myosin in the biopsies was taken into account. Data for wild-type myosin of slow soleus muscle fibers support previous findings that for the slow myosin isoform S( *) and F( *) are significantly lower than for fast myosin e.g., of rabbit psoas muscle. The data indicate that two mutations, R723G and R719W, are associated with an increase in resistance to elastic distortion of the individual mutated myosin heads whereas mutation I736T has essentially no effect. The data strongly support the notion that major elastic distortion occurs within the converter itself. Apparently, the compliance depends on specific residues, e.g., R719 and R723, presumably located at strategic positions near the long alpha-helix of the light chain binding domain. Because amino acids 719 and 723 are nonconserved residues, cross-bridge stiffness may well be specifically tuned for different myosins.

Engineered mutants of HGF/SF with reduced binding to heparan sulphate proteoglycans, decreased clearance and enhanced activity in vivo.

BACKGROUND: Although a number of growth factors bind cell-surface heparan sulphate proteoglycans (HSPGs), the role of this interaction is unclear except for fibroblast growth factor which requires HSPG binding for signalling. Hepatocyte growth factor/scatter factor (HGF/SF) plays important roles in mammalian development and tissue regeneration and acts on target cells through a specific receptor tyrosine kinase encoded by the c-met proto-oncogene. This factor also binds HSPGs with high affinity, but conflicting data have been reported on the role of HSPG binding in HGF/SF signalling. RESULTS: To map the binding sites for HSPG and the Met receptor in HGF/SF, we have engineered a number of HGF/SF mutants in which several clusters of solvent-accessible residues in the hairpin structure of the amino-terminal domain or in kringle 2 have been replaced. Two of the mutants (HP1 and HP2) showed greatly decreased (more than 50-fold) affinity for heparin and HSPGs but retained full mitogenic and motogenic activities on target cells in culture. Furthermore, when compared with wild-type HGF/SF, the HP1 mutant exhibited a delayed clearance from the blood, higher tissue levels and a higher induction of DNA synthesis in normal, adult murine liver. CONCLUSIONS: These results establish the following: the binding sites in HGF/SF for Met and for HSPGs can be dissociated by protein engineering; high-affinity binding of HGF/SF to HSPGs is not essential for signalling; one role of HSPG binding in the HGF/SF system appears to be sequestration and degradation of the growth factor; and HGF/SF mutants with decreased affinity for HSPGs exhibit enhanced activity in vivo.

A role of the cryptic gene in the correct establishment of the left-right axis.

During vertebrate embryogenesis, a left-right axis is established. The heart, associated vessels and inner organs adopt asymmetric spatial arrangements and morphologies. Secreted growth factors of the TGF-beta family, including nodal, lefty-1 and lefty-2, play crucial roles in establishing left-right asymmetries [1] [2] [3]. In zebrafish, nodal signalling requires the presence of one-eyed pinhead (oep), a member of the EGF-CFC family of membrane-associated proteins [4]. We have generated a mutant allele of cryptic, a mouse EGF-CFC gene [5]. Homozygous cryptic mutants developed to birth, but the majority died during the first week of life because of complex cardiac malformations such as malpositioning of the great arteries, and atrial-ventricular septal defects. Moreover, laterality defects, including right isomerism of the lungs, right or left positioning of the stomach and splenic hypoplasia were observed. Nodal gene expression in the node was initiated in cryptic mutant mice, but neither nodal, lefty-2 nor Pitx2 were expressed in the left lateral plate mesoderm. The laterality defects observed in cryptic(-/-) mice resemble those of mice lacking the type IIB activin receptor or the homeobox-containing factor Pitx2 [6] [7] [8] [9], and are reminiscent of the human asplenic syndrome [10]. Our results provide genetic evidence for a role of cryptic in the signalling cascade that determines left-right asymmetry.

Intrapleural fibrinolytic treatment of traumatic clotted hemothorax.

STUDY OBJECTIVE: To evaluate the role of intrapleural fibrinolytic treatment (IPFT) in traumatic clotted hemothorax. DESIGN AND PATIENTS: Between August 1995 and February 1997, 24 patients with traumatic clotted hemothorax were included. Streptokinase (SK), 250,000 IU, or urokinase (UK), 100,000 IU, diluted in 100 mL of saline solution was given daily. We administered 5.0+/-1.8 (range, 2 to 9) doses of SK or 6.25+/-5.97 (range, 2 to 15) doses of UK. SETTING: Dicle University School of Medicine, Thoracic and Cardiovascular Surgery Department. RESULTS: Complete response, which was defined as resolution of symptoms with complete drainage of fluid and no residual space radiographically, occurred in 15 (62.5%) patients. Partial response, which was defined as resolution of symptoms with a small pleural cavity, occurred in seven (29.2%) patients. Two patients (8.3%) required decortication; they were defined as nonresponders. The mean period of time between the diagnosis and fibrinolytic treatment (FT) was 11.65+/-6.38 (range, 4 to 25) days. There were no complications related to IPFT. There was no mortality during the course of IPFT. CONCLUSION: The use of intrapleural fibrinolytic agents has resulted in resolution of clotted hemothorax with an overall success rate of 91.7%. We recommend that IPFT should be added to the algorithm for management of clotted hemothorax before proceeding with minithoracotomy or pleural decortication.

Penetrating chest injuries in children: a review of 94 cases.

Ninety-four children with penetrating chest injuries were treated at Dicle University School of Medicine during a 6-year period. The mean age was 11.51 +/- 3.31 years, and the male:female ratio was 5.25:1. Forty-five had stab wounds, 27 had high-velocity gunshot wounds, 13 had low-velocity gunshot wounds, seven had a bomb (shrapnel) injury, one had a shotgun wound, and one had a horse bite. Sixty patients had isolated thoracic injuries, and 34 had associated injuries. The most common thoracic injury was hemothorax (28), followed by hemopneumothorax (25). Tube thoracostomy alone was sufficient in 79.8% of the patients (75 of 94). Thoracotomy was performed in 4.25% (4 of 94). In two of the five observed patients, delayed hemothorax developed. The mean duration of hospitalization was 5.13 +/- 1.93 days. The mean Injury Severity Score was 14.71 +/- 8.62. Prophylactic antibiotics were used in all patients. The morbidity rate was 8.51% (8 of 94). Only one death occurred after cervical tracheal repair. The study suggests that the majority of penetrating chest injuries in children can be treated successfully by tube thoracostomy alone or in conjunction with expectant observation.

Surgical treatment of pulmonary hydatidosis in children: experience in 92 patients.

Ninety-two patients with a total of 112 pulmonary hydatid cysts underwent surgical treatment in our clinic between January 1980 and January 1992. Nine patients were found to have concomitant liver and pulmonary hydatid cysts. There were 65 boys and 27 girls; the age range was 5 to 14 years. The authors performed cystotomy and capitonnage in 78 patients, lobectomy in 9, segmentectomy in 3, and wedge resection in 2. Of 9 cases with concomitant liver and pulmonary hydatid cysts, 8 were approached by right thoracophrenotomy and 1 by median sternotomy and right phrenotomy. A one-stage operation via median sternotomy was performed in 3 patients who had bilateral pulmonary hydatid cysts. To prevent the development of secondary hydatid cysts because of spillage of hydatid fluid, the authors used 10% povidone-iodine poly (1-vinyl-2-pyrrolidone)-iodine complex intraoperatively as a scolicidal agent. There was no mortality or recurrence in the series.

Thoracic trauma in the elderly.

Thoracic trauma in the elderly population constitutes a major challenge for both thoracic and trauma surgeons as their presentation and outcomes differ from the adult population in addition to their high morbidity and mortality. One hundred and one patients, 60 years of age or older, with thoracic trauma were treated at Dicle University School of Medicine during a 6-year period. Eighty-five per cent were male and 15% were female with a mean age of 64.5 years. The cause of thoracic injury was blunt in 77.2% and penetrating in 22.8% of the patients. Sixty-two patients (61.4%) had isolated thoracic injuries. The median Injury Severity Score (ISS) was 23. The morbidity rate was 23.8%. The mortality rate was 16.8%. Seven of 10 patients (70%) who had an ISS greater than 25 died, whereas six of 24 (25%) patients with an ISS between 17 and 25, and four of 67 (5.9%) patients with an ISS less than 16 died. In the elderly the morbidity and mortality rates were higher for blunt trauma compared with penetrating trauma. For ISS greater than 25 the mortality rate was 71.4% for blunt and 66.6% for penetrating trauma. As the morbidity and mortality rate are significantly higher in the elderly patients the approach to these patients should include recognition of their high risk for morbidity and mortality, especially for those who had an ISS greater than 25.

Severe enamel abrasion due to misuse of an air polishing device.

OBJECTIVES: In this case report, a 28-year-old male patient who severely injured the enamel tissue of his upper incisors due to excessive self-performed air polishing has been presented. Air polishing devices are frequently used in initial or supportive periodontal therapy to remove supragingival plaque and dental stains. Enamel tissue is minimally affected by air polishing when appropriately performed by a professional. However, excessive air polishing may have detrimental effects even on the intact enamel. The aim of this case report is to present a patient who severely injured the enamel surfaces of his upper incisors due to excessive self-performed air polishing. METHODS: A case of severe enamel abrasions in a 28-year-old male patient who injured the enamel surfaces of his upper incisors following several self-performed air polishing sessions has been presented. RESULTS: Severely abraded enamel surfaces of the upper incisors were present and during the course of therapy restored by composite restorations to establish a satisfactory clinical appearance. CONCLUSIONS: In clinical practice, air polishing can be performed rather safely on intact enamel and is a beneficial procedure in initial and supportive periodontal therapy when performed by a professional under recommended operating conditions. However, review of the literature reveals that air polishing may be harmful on tooth and surrounding structures unless carried out cautiously. Furthermore, excessive use of air polishing devices, especially by unauthorized personnel may be damaging and lead to severe abrasion of enamel tissue.

Near-total esophageal exclusion in the treatment of late-diagnosed esophageal perforation.

Two cases of late-diagnosed esophageal perforation were successfully treated with near-total esophageal exclusion, using cervical T-tube esophagostomy with circumferential suture fixation of the lower arm of the T-tube. Esophageal stricture developed at the site of catgut or dexon tie. This complication can be managed with esophageal dilation. Drainage-tube gastrostomy proved to be unnecessary.

Angiotensin-converting enzyme inhibition, autonomic activity, and hemodynamics in patients with heart failure who perform isometric exercise.

Effects of angiotensin-converting enzyme inhibition (ACEI) on autonomic responses and hemodynamics in patients with congestive heart failure (CHF) subjected to isometric exercise have not been studied. We tested whether acute ACEI might influence the effects of isometric exercise in patients with CHF. In the first part of the study we showed that isometric exercise increased blood pressure in the control group and in the CHF group, whereas cardiac output increased only in the control group. Stroke volume remained unchanged in the control group, whereas it decreased significantly in CHF group. We next analyzed the effect of acute ACEI (5 mg ramipril) on the decrease in cardiac output during isometric stress in patients with CHF. During isometric exercise mean blood pressure and heart rate increased similarly in both groups. However, cardiac output decreased during placebo by -0.48 +/- 0.12 L/min (p < 0.01) but not during ACEI. Spectral analysis of blood pressure showed an increase (p < 0.01) in the high-frequency parasympathetic component from 7.3% +/- 3.6% to 18.1% +/- 9.5% after ACEI. norepinephrine plasma levels increased after isometric stress in the placebo group, whereas other hormones did not change. ACEI prevented the norepinephrine increase after isometric stress. Thus the decrease in cardiac output during isometric exercise in patients with CHF was prevented by acute ACEI. The effect of ACE inhibition may be related to reduced sympathetic activity.

[Penetrating heart wounds. An analysis of 29 cases]. / Penetran kalp yaralanmali olgularimizin analizi.

Twenty-nine cases, penetrating wounds to the heart, who were operated in the thoracic and cardiovascular surgery department of our University during January 1995-August 2000 were reevaluated. Twenty nine patients were men and the mean age was 22.6 (min 12, max 45). The cause of injury was stab wounds in 28 cases and gunshot wounds in 1 case. Besides the clinical findings; teleradiography, echocardiography were valuable for the diagnosis. Surgery was applied in emergent conditions for all cases. Interventions were performed 28 anterolateral thoracotomy, 1 bilaterally anterolateral thoracotomy and additionally 4 laparatomy were performed. The most frequent side of injury was right ventricule. Our mortality ratio was 17.2%, morbidity ratio 17.2%. We think that fast transportation, urgent diagnostic study and immediate surgical intervention important parameters which decrease of the mortality.

Traumatic oesophageal perforation.

Sixteen patients were treated for traumatic oesophageal perforation (13 cervical, 3 thoracic) over a 16-year period. In 14 cases the trauma was penetrating. The median delay from injury to treatment was 32 hours and the mean period of hospitalization was 26 days. The treatment procedures were two-layer primary closure with or without drainage, drainage alone and near-total oesophageal exclusion with cervical T-tube oesophagostomy. Postoperative complications were cervical oesophageal leak in two patients and tracheo-oesophageal fistula and oesophageal stenosis, each in one case. Of the eight patients treated within 24 hours of perforation, two died, and of the eight treated later, four died (overall mortality 37.5%). The heightened mortality after delayed diagnosis illustrates the prognostic importance of a high index of suspicion. To prevent leakage, buttressing with viable tissue following primary closure can be useful, especially after delayed diagnosis. Because of the continuing controversy concerning management of late-diagnosed oesophageal perforation, individualized treatment is widely advocated.

Penetrating chest injuries: unusually high incidence of high-velocity gunshot wounds in civilian practice.

Penetrating chest injuries are a challenge to the thoracic or trauma surgeon. Penetrating thoracic trauma, especially that due to high-velocity gunshot wounds, is increasing at an alarming rate in our region. We report our experience with penetrating chest injuries mainly due to high-velocity gunshot wounds. During a period of 6 years we retrospectively reviewed the hospital records of 755 patients admitted to the Department of Thoracic and Cardiovascular Surgery, Dicle University School of Medicine, with the diagnosis of penetrating thoracic trauma. The mean age was 27.48 years, and 89-8% were male. The causes of penetrating injury were stab wounds in 45.3% and gunshot wounds in 54.7%. About 30% of the wounds were due to high-velocity gunshots; and among the gunshot wounds 56.2% were due to high-velocity shots. The most common thoracic injury was hemothorax (n = 190) followed by hemopneumothorax (n = 184). Isolated thoracic injuries were found in 53% of the patients. Nonoperative management was sufficient in 92% of the patients. Thoracotomy was performed in 8.1%. The mean duration of hospitalization was 11.2 days. The mean injury severity score (ISS) was 20.17 +/- 13.87. The morbidity was 23.3% and the mortality 5.6%. Fifty percent of all deaths were due to adult respiratory distress syndrome. Altogether 17% of patients with an ISS > 25 died, whereas only 0.9% of those with a score < 16 died. The mortality due to firearms was 8.95%. We concluded that in civilian practice chest tube thoracostomy remains by far the most common method of treating penetrating injury to the chest. The easy availability of high-velocity guns will continue to increase the number of civilians injured by these weapons.