Mutant G protein alpha subunit activated by Gbeta gamma: a model for receptor activation?

How receptors catalyze exchange of GTP for GDP bound to the Galpha subunit of trimeric G proteins is not known. One proposal is that the receptor uses the G protein's betagamma heterodimer as a lever, tilting it to pull open the guanine nucleotide binding pocket of Galpha. To test this possibility, we designed a mutant Galpha that would bind to betagamma in the tilted conformation. To do so, we excised a helical turn (four residues) from the N-terminal region of alpha(s), the alpha subunit of G(S), the stimulatory regulator of adenylyl cyclase. In the presence, but not in the absence, of transiently expressed beta(1) and gamma(2), this mutant (alpha(s)Delta), markedly stimulated cAMP accumulation. This effect depended on the ability of the coexpressed beta protein to interact normally with the lip of the nucleotide binding pocket of alpha(s)Delta. We substituted alanine for an aspartate in beta(1) that binds to a lysine (K206) in the lip of the alpha subunit's nucleotide binding pocket. Coexpressed with alpha(s)Delta and gamma(2), this mutant, beta(1)-D228A, elevated cAMP much less than did beta(1)-wild type; it did bind to alpha(s)Delta normally, however, as indicated by its unimpaired ability to target alpha(s)Delta to the plasma membrane. We conclude that betagamma can activate alpha(s) and that this effect probably involves both a tilt of betagamma relative to alpha(s) and interaction of beta with the lip of the nucleotide binding pocket. We speculate that receptors use a similar mechanism to activate trimeric G proteins.     

A physical basis for protein secondary structure

A physical theory of protein secondary structure is proposed and tested by performing exceedingly simple Monte Carlo simulations. In essence, secondary structure propensities are predominantly a consequence of two competing local effects, one favoring hydrogen bond formation in helices and turns, the other opposing the attendant reduction in sidechain conformational entropy on helix and turn formation. These sequence specific biases are densely dispersed throughout the unfolded polypeptide chain, where they serve to preorganize the folding process and largely, but imperfectly, anticipate the native secondary structure.     

Calibration and optimization of 3D digital breast tomosynthesis guided near infrared spectral tomography

Calibration of a three-dimensional multimodal digital breast tomosynthesis (DBT) x-ray and non-fiber based near infrared spectral tomography (NIRST) system is challenging but essential for clinical studies. Phantom imaging results yielded linear contrast recovery of total hemoglobin (HbT) concentration for cylindrical inclusions of 15 mm, 10 mm and 7 mm with a 3.5% decrease in the HbT estimate for each 1 cm increase in inclusion depth. A clinical exam of a patient’s breast containing both benign and malignant lesions was successfully imaged, with greater HbT was found in the malignancy relative to the benign abnormality and fibroglandular regions (11 μM vs. 9.5 μM). Tools developed improved imaging system characterization and optimization of signal quality, which will ultimately improve patient selection and subsequent clinical trial results.OCIS codes: (120.3890) Medical optics instrumentation, (170.0110) Imaging systems, (170.3830) Mammography     

Compression screw fixation without bone grafting for scaphoid fibrous nonunion

Purpose

Bone graft is often recommended as an adjuvant for treatment of scaphoid nonunions. However, recent literature has suggested that fibrous nonunion may be suited to treatment with rigid fixation without bone grafting. This work reported on outcomes of compression screw fixation for established scaphoid fibrous nonunions without bone graft.

Methods

Fourteen patients underwent surgical compression screw fixation without bone grafting of scaphoid fibrous nonunion between January 1, 2000, and December 31, 2012, with minimum follow-up until the time of clinical and radiographic healing. Fibrous nonunion of the scaphoid was defined as a scaphoid fracture with all of the following features: (1) persistent tenderness, (2) incomplete trabecular bridging on three X-ray views, (3) injury that had occurred at a minimum of 6 months prior to surgery, and (4) identification of fibrous union at the time of surgery. Outcomes were assessed with range of motion assessment, Disability of the Arm, Shoulder and Hand (DASH) scores, and plain radiographs.

Results

Twelve of the 14 patients healed successfully, while two patients required secondary vascularized bone grafting. Both unhealed patients sustained proximal pole fractures and had a duration of ≥1 year from injury to surgery. Average time to healing was 4.4 ± 2.0 months. Average flexion was 73 ± 22° and average extension was 66 ± 22° postoperatively. Average grip strength was 90 ± 25 lbs on the operative side. Mean postoperative pain score was 1.4 (range, 0 to 7). Mean postoperative DASH score was 10.2 (range, 0 to 52). Increasing age and an interval from injury to surgery of >1 year correlated with worse DASH and pain scores.

Conclusions

Patients with fibrous scaphoid nonunion demonstrated good results with rigid fixation without bone grafting. Increasing age and >1-year interval between injury and surgery resulted in lower self-assessed outcomes.

Level of Evidence

Therapeutic Level IV, retrospective case series     

Risk factors in early life as predictors of adult heart disease: the Bogalusa Heart Study

The adult heart diseases, coronary artery disease and essential hypertension, are now clearly recognized to begin in childhood. The evidence comes from autopsy studies of cardiovascular-renal changes in the first two decades of life. Cardiovascular risk factors can be identified in children just as in adults and these have a high correlation with the anatomic disease. This relationship underscores the importance of risk factor screening of children. Of interest is that clinical risk factors tend to persist within a rank (track) so that studies in childhood can be predictive of future levels. Behavior and lifestyle of eating, cigarette smoking, alcohol intake, and use of oral contraceptive pills influence risk factors in children. Familial aggregation of risk factors are also noted. Studies of apolipoproteins, B and A-I, have identified subsets of children that have a greater frequency of paternal myocardial infarction. The findings from the Bogalusa Heart Study and other epidemiological studies of children show the need to begin prevention of adult heart disease in early life. Approaches to prevention should include high risk families and children and a public health or population approach. Cardiovascular health education for elementary school children should be directed to children in the general population in an effort to encourage them to adopt healthy life styles.     

Metabolic programming in the immediate postnatal period

In recent decades, there has been a dramatic increase in the incidence of obesity in all age groups of the population in the USA. In addition to genetics and life style changes, the important role of metabolic programming effects in the etiology of the obesity epidemic is being increasingly recognized. Although the role of a compromised intrauterine environment in fetal metabolic programming is well documented to contribute to the development of adult-onset diseases, vulnerability in the immediate postnatal period to similar conditions has also been shown. Metabolic programming effects induced by altered nutritional experiences in the immediate postnatal period can give rise to long-term consequences in the context of the current obesity epidemic.     

Early activation of mucosal dendritic cells and macrophages in acute Campylobacter colitis and cholera: An in vivo study

Background and Aim: Macrophages and dendritic cells are closely related mononuclear phagocytic cells. Little is known about their in vivo role in acute intestinal bacterial infections in humans. We undertook to evaluate these cells in rectal mucosal biopsies of patients with acute colitis. Methods: All mucosal mononuclear phagocytic cells in rectal biopsies of patients with acute Campylobacter colitis (n = 5), shigellosis (n = 5), and cholera (n = 10) were evaluated ultrastructurally and compared with those in controls (n = 5). Results: Mononuclear phagocytic cells in the superficial rectal mucosa showed a higher prevalence of ultrastructural features of activation in Campylobacter colitis and cholera than in controls. A lower prevalence of features of activation with increased monocytes was seen in shigellosis. Cells with the ultrastructural morphology of activated dendritic cells constituted 41% and 45% of all mononuclear phagocytic cells in two of five patients with Campylobacter colitis and 4–22% of cells in four of 10 patients with cholera. Their presence in patients with Campylobacter colitis was associated with significant surface epithelial damage and prominent acute inflammatory changes in the mucosa. Conclusions: This is the first ultrastructural study to show activated macrophages and dendritic cells in vivo in acute Campylobacter colitis and cholera. Dendritic cell activation occurred early in the clinical course of these infections. Surface epithelial damage may play a role in the activation of dendritic cells.     

Nodal Langerhans cell histiocytosis with prominent eosinophilic emperipolesis

Multinucleated giant cells of osteoclastic type are known to occur in nodal and extranodal lesions of Langerhans cell histiocytosis (LCH). These giant cells are thought to be derived from phagocytic histiocytes, which correlate with the degree of necrosis in LCH lesions. Emperipolesis commonly seen in Rosai–Dorfman disease is a distinct phenomenon characterized by intact phagocytosed cells in an intracytoplasmic vacuole protected from proteolytic digestion. We present a case of emperipolesis of inflammatory cells especially of eosinophils by multinucleated giant cells of Langhans type in a lymph node involved by LCH—a finding that has not been described previously in the literature. Diagn. Cytopathol. 2015;43:1000–1002. © 2015 Wiley Periodicals, Inc.