Simulating backspatter of blood from cranial gunshot wounds using pig models.

Few studies have examined the biomechanical basis for backspatter from cranial gunshot wounds. Backspatter is material which travels against the direction of fire following ejection from a gunshot entrance wound. Our paper focuses on the use of animals for reconstructing this phenomenon. Five live pigs and several slaughtered pigs were shot using either 9 × 19 mm, 115 grain, full metal jacketed ammunition or .22 long rifle, 40 grain, lead, round-nose ammunition. A high-speed camera was used to record the entrance wound formation and backspatter. A small amount of backspattered material was produced with all targets, and blood backspatter was seen in a few cases. However, we conclude that our model provides an understanding of the phenomenon of backspatter and the physical mechanisms associated with it. The various components of the mechanism of backspatter formation are complex and overlap. The principle mechanism observed in pig cranial gunshots was the high-speed impact response of the skin overlying the skull bone. This study has also produced evidence supporting the view that backspatter can result from the splashing of superficial blood if it is already present on the skin. Subcutaneous gas effects have been demonstrated for backspatter from contact shots. There has been no clear evidence of the role of the collapse of a temporary cavity within the brain.

Effectiveness of the STOPP/START (Screening Tool of Older Persons' potentially inappropriate Prescriptions/Screening Tool to Alert doctors to the Right Treatment) criteria: systematic review and meta-analysis of randomized controlled studies.

WHAT IS KNOWN AND OBJECTIVE: STOPP/START are explicit screening tools that identify potentially inappropriate prescribing in older adults. Our objective was to update our 2013 systematic review that showed limited evidence of impact, using new evidence from randomized controlled trials (RCTs) assessing clinical, humanistic and economic outcomes in older adults. METHODS: We performed a search of PubMed, EMBASE, CINAHL, Web of Science and grey literature for RCTs published in English since the previous review through June 2014. The Cochrane Risk of Bias Tool was used. We performed a meta-analysis on the effect of STOPP on potentially inappropriate medication (PIM) rates and a narrative synthesis on other outcomes. RESULTS AND DISCUSSION: Four RCTs (n = 1925 adults) from four countries were included, reporting both acute (n = 2) and long-term care (n = 2) patients. Studies differed in implementation. Two studies were judged to have low risk, and two to have moderate-to-high risk of bias in key domains. Meta-analysis found that the STOPP criteria reduced PIM rates in all four studies, but study heterogeneity (I(2) = 86·7%) prevented the calculation of a meaningful statistical summary. We found evidence that use of the criteria reduces falls, delirium episodes, hospital length-of-stay, care visits (primary and emergency) and medication costs, but no evidence of improvements in quality of life or mortality. WHAT IS NEW AND CONCLUSION: STOPP/START may be effective in improving prescribing quality, clinical, humanistic and economic outcomes. Additional research investigating these tools is needed, especially in frail elderly and community-living patients receiving primary care.

Probing configuration mixing in 12Be with Gamow-Teller transition strengths.

We present a novel technique for studying the quenching of shell gaps in exotic isotopes. The method is based on extracting Gamow-Teller (ΔL=0, ΔS=1) transition strengths [B(GT)] to low-lying states from charge-exchange reactions at intermediate beam energies. These Gamow-Teller strengths are very sensitive to configuration mixing between cross-shell orbitals, and this technique thus provides an important complement to other tools currently used to study cross-shell mixing. This work focuses on the N=8 shell gap. We populated the ground and 2.24 MeV 0+ states in 12Be using the 12B(1+) (7Li, 7Be) reaction at 80 MeV/u in inverse kinematics. Using the ground-state B(GT) value from ß-decay measurements (0.184±0.007) as a calibration, the B(GT) for the transition to the second 0+ state was determined to be 0.214±0.051. Comparing the extracted Gamow-Teller strengths with shell-model calculations, it was determined that the wave functions of the first and second 0+ states in 12Be are composed of 25±5% and 60±5% (0s)4(0p)8 configurations, respectively.

Enhanced quadrupole collectivity at N = 40: the case of neutron-rich Fe isotopes.

The transition rates for the 2(1)+ states in (62,64,66)Fe were studied using the recoil distance Doppler-shift technique applied to projectile Coulomb excitation reactions. The deduced E2 strengths illustrate the enhanced collectivity of the neutron-rich Fe isotopes up to N = 40. The results are interpreted using the generalized concept of valence proton symmetry which describes the evolution of nuclear structure around N = 40 as governed by the number of valence protons with respect to Z ≈ 30. The trend of collectivity suggested by the experimental data is described by state-of-the-art shell-model calculations with a new effective interaction developed for the fpgd valence space.

Lifetime measurement of the 2(1)+ state in 20C.

Establishing how and when large N/Z values require modified or new theoretical tools is a major quest in nuclear physics. Here we report the first measurement of the lifetime of the 2(1)+ state in the near-dripline nucleus 20C. The deduced value of τ(2(1)+)=9.8±2.8(stat)(-1.1)(+0.5)(syst) ps gives a reduced transition probability of B(E2; 2(1)+→0(g.s.)+)=7.5(-1.7)(+3.0)(stat)(-0.4)(+1.0)(syst) e2 fm4 in good agreement with a shell model calculation using isospin-dependent effective charges.

Tobramycin pharmacokinetics in patients with cystic fibrosis before and after bilateral lung transplantation.

STUDY OBJECTIVES: To compare the pharmacokinetics (PK) of tobramycin in patients with cystic fibrosis (CF) before and after bilateral lung transplantation, in order to evaluate optimal dosing practices post transplant. DESIGN: Retrospective, single-center, chart review study, in which tobramycin concentrations from CF patients were used to calculate PK parameters, including elimination rate constant, half-life, volume of distribution (Vd), area under the curve (AUC), and clearance before and after lung transplantation. SETTING: Medical school-affiliated teaching hospital. PATIENTS: Eight patients with CF, who received a bilateral lung transplant from January 1, 2005 through August 1, 2009 (4 males, 4 females; mean age 26.3 years). INTERVENTIONS: None. MAIN RESULTS: Sixty-nine sets of pre- (n=52) and post transplant (n=17) tobramycin concentrations were available. PK parameters were significantly altered post transplant. Elimination rate constant decreased 38% from 0.26±0.1 to 0.16±0.1 h(-1) (P<0.001), with a related increase of 200% in half-life from 2.8±0.8 to 8.4±8.7 h (P<0.001). Clearance decreased 25% post transplant from 67.3±32.3 to 50.2±15.9 mL/min (P=0.04). No statistically significant change occurred in AUC or Vd after transplant, although a trend was seen toward increased Vd. Dosage requirements after transplantation were significantly lower, 10.7±2.5 and 7.6±1.6 mg/kg/day, pre and post transplant, respectively (P<0.001). Concentrations were also evaluated in 2 time periods: 0-3 weeks and ≥6 weeks post transplant, based on available data. Clearance and Vd ≥6 weeks post transplant did not significantly differ from pre-transplant values (P=0.28 and 0.54, respectively), suggesting that these changes may be temporary. CONCLUSIONS: The results suggest that tobramycin PK are altered in patients with CF after bilateral lung transplantation, although no clear trend was seen owing to inter-patient variability. We propose that PK parameters should be reassessed during each treatment course post transplant.

Murine tissue amyloid protein AA. NH2-terminal sequence identity with only one of two serum amyloid protein (ApoSAA) gene products.

Amyloid protein AA is the presumptive fragment of an acute phase serum apolipoprotein, apoSAA. Two major murine apoSAA isotypes (apoSAA1 and apoSAA2) have been identified. The NH2-terminal amino acid sequences of purified murine apoSAA1 and apoSAA2 have been examined and compared with that of murine amyloid protein AA. Our results indicate that apoSAA1 and apoSAA2 are separate gene products and that amyloid protein AA has NH2-terminal amino acid sequence identity with only one of these isotypes, namely apoSAA2.

Effects of external influences on synthetic cannabinoid trends in New Zealand, 2014 to 2020.

Over the past decade, synthetic cannabinoids have inundated the global market and now form the largest category of new psychoactive substances. Once these chemicals are available on the global market, they can be applied to plant material in a clandestine environment to create an end-product that is smoked by the user. The synthetic cannabinoids AMB-FUBINACA and 5F-ADB were most frequently detected between 2017 and the beginning of 2019. More recently, these two appear to have been replaced by different synthetic cannabinoids. This investigation summarises the recent trends in synthetic cannabinoids detected in New Zealand between 2017 and 2020 and outlines the potential factors influencing these trends.

Identification of two intermediates during processing of profilaggrin to filaggrin in neonatal mouse epidermis.

A major event in the keratinization of epidermis is the production of the histidine-rich protein filaggrin (26,000 mol wt) from its high molecular weight (greater than 350,000) phosphorylated precursor (profilaggrin). We have identified two nonphosphorylated intermediates (60,000 and 90,000 mol wt) in NaSCN extracts of epidermis from C57/Bl6 mice by in vivo pulse-chase studies. Results of peptide mapping using a two-dimensional technique suggest that these intermediates consist of either two or three copies of filaggrin domains. Each of the intermediates has been purified. The ratios of amino acids in the purified components are unusual and essentially identical. The data are discussed in terms of a precursor containing tandem repeats of similar domains. In vivo pulse-chase experiments demonstrate that the processing of the high molecular weight phosphorylated precursor involves dephosphorylation and proteolytic steps through three-domain and two-domain intermediates to filaggrin. These processing steps appear to occur as the cell goes through the transition cell stage to form a cornified cell.

Evolution of structure and function of proteases.

One of the striking features of the proteolytic enzymes as a group is the immense variety of biological functions served by enzymes employing one of a few basic mechanisms. For example, in the higher animals, enzymes for activation of zymogens (trypsin), for digestion of dietary proteins (trypsin, chymotrypsin, elastase), for blood clotting (thrombin), for clot lysis (plasmin), and for sensing pain (kallikrein) all appear to use the same mechanism and to have evolved from the same ancestral gene by the process of gene duplication and subsequent divergent evolution. Equally striking is the variety of chemical solutions of the same functional problem, such as the peptide-bond cleavage by sulfhydryl proteases on the one hand and serine proteases on the other.

Recoverin: a calcium sensitive activator of retinal rod guanylate cyclase.

Vertebrate retinal photoreceptors recover from photoexcitation-induced hydrolysis of guanosine 3', 5'-monophosphate (cyclic GMP) by resynthesizing cyclic GMP, which reopens cation channels that have been closed by light. Activation of guanylate cyclase by light-induced depletion of cytosolic calcium is a key event in this recovery process. This cyclase has now been shown to be regulated by a 23-kilodalton calcium binding protein. The protein is present in both rod and cone photoreceptors and was named recoverin because it promotes recovery of the dark state. The amino acid sequence of recoverin exhibits three potential calcium binding sites (EF hands). That recoverin binds calcium was confirmed with calcium-45 and by observing calcium-induced changes in its tryptophan fluorescence. Recoverin activated guanylate cyclase when free calcium was lowered from 450 to 40 nM, an effect that was blocked by an antibody to recoverin. Thus, guanylate cyclase in retinal rods is stimulated during recovery by the calcium-free form of recoverin. A comparison of recoverin with other calcium binding proteins reveals that it may represent, along with the protein visinin, a family of proteins that are regulated by submicromolar calcium concentrations.

The identification and quantification of synthetic cannabinoids seized in New Zealand in 2017.

There has been an explosion of new psychoactive substances (NPS) over the past decade, with synthetic cannabinoids comprising one of the more extensive and chemically diverse groups. Synthetic cannabinoids, like other NPS, are continually evolving with slight alterations in chemical structure, which can lead to unintended and harmful effects for the user. Furthermore, the clandestine preparation of plant material containing one or more synthetic cannabinoid can result in an unevenly distributed product, which poses an additional risk to the user of increased doses. This investigation aimed at providing a snapshot of synthetic cannabinoids in New Zealand in 2017, including the concentrations of synthetic cannabinoids in plant material. Overall, ten different synthetic cannabinoids were detected, with AMB-FUBINACA and 5F-ADB comprising the majority of samples analysed. The synthetic cannabinoid AMB-FUBINACA displayed the greatest range of concentration in plant material, from 5 to over 400 g of synthetic cannabinoid per kilogram of plant material. There was also geographical variation in the synthetic cannabinoids depending on where in New Zealand they were seized from.

Protein-tyrosine phosphatase activity of CD45 is activated by sequential phosphorylation by two kinases.

We describe a potential regulatory mechanism for the transmembrane protein-tyrosine phosphatase CD45. Phosphorylation on both tyrosine and serine residues in vitro results in an activation of CD45 specifically toward one artificial substrate but not another. The activation of these kinases appears to be order dependent, as it is enhanced when phosphorylation of tyrosine precedes that of serine but phosphorylation in the reverse order yields no activation. Any of four protein-tyrosine kinases tested, in combination with the protein-serine/threonine kinase, casein kinase II, was capable of mediating this activation in vitro. The time course of phosphorylation of CD45 in response to T-cell activation is consistent with the possibility that this regulatory mechanism is utilized in vivo.

Mass spectrometric measurement of protein amide hydrogen exchange rates of apo- and holo-myoglobin.

Measurement of backbone amide hydrogen exchange rates can provide detailed information concerning protein structure, dynamics, and interactions. Although nuclear magnetic resonance is typically used to provide these data, its use is restricted to lower molecular weight proteins that are soluble at millimolar concentrations. Not subject to these limitations is a mass spectrometric approach for measuring deuterium incorporation into proteins that are subsequently proteolyzed by pepsin; the resulting peptide masses are measured using a flowing-fast atom bombardment ionization source (Zhang Z, Smith DL, 1993, Protein Sci 2:522-531). In the current study, amide deuterium incorporation for intact apo- and holo-myoglobin was measured using liquid chromatography coupled directly to an electrospray ionization (LC/MS) source. Electrospray ionization provided a more complete coverage of the protein sequence and permitted the measurement of deuterium incorporation into intact proteins. Tandem mass spectrometry was used to rapidly identify the peptic peptides. It was found that within 30 s, the amides in apo-myoglobin were 47% deuterated, whereas holo-myoglobin was 12% deuterated. Peptic digestion and LC/MS demonstrated that regions represented by peptic peptides encompassing positions 1-7, 12-29, and 110-134 were not significantly altered by removal of the heme. Likewise, destabilized regions were identified within positions 33-106 and 138-153.

Sequence analysis of peptide mixtures by automated integration of Edman and mass spectrometric data.

A computer algorithm is described that utilizes both Edman and mass spectrometric data for simultaneous determination of the amino acid sequences of several peptides in a mixture. Gas phase sequencing of a peptide mixture results in a list of observed amino acids for each cycle of Edman degradation, which by itself may not be informative and typically requires reanalysis following additional chromatographic steps. Tandem mass spectrometry, on the other hand, has a proven ability to analyze sequences of peptides present in mixtures. However, mass spectrometric data may lack a complete set of sequence-defining fragment ions, so that more than one possible sequence may account for the observed fragment ions. A combination of the two types of data reduces the ambiguity inherent in each. The algorithm first utilizes the Edman data to determine all hypothetical sequences with a calculated mass equal to the observed mass of one of the peptides present in the mixture. These sequences are then assigned figures of merit according to how well each of them accounts for the fragment ions in the tandem mass spectrum of that peptide. The program was tested on tryptic and chymotryptic peptides from hen lysozyme, and the results are compared with those of another computer program that uses only mass spectral data for peptide sequencing. In order to assess the utility of this method the program is tested using simulated mixtures of varying complexity and tandem mass spectra of varying quality.

Beta-methylthio-aspartic acid: identification of a novel posttranslational modification in ribosomal protein S12 from Escherichia coli.

Utilizing microscale chemical derivatization reactions and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, we have identified a novel posttranslational modification of aspartic acid, beta-methylthio-aspartic acid. The modified residue is located at position 88 in ribosomal protein S12 from Escherichia coli, a phylogenetically conserved protein that has been implicated in maintaining translational accuracy of the ribosome.

Direct sequence analysis of proteins by in-source fragmentation during delayed ion extraction.

Continuous segments of amino acid sequence information as long as 41 residues have been deduced by interpretation of matrix-assisted laser desorption/ionization-generated ion signals dominated by Cn fragmentation within the ion source of a linear time-of-flight mass spectrometer utilizing delayed ion extraction. The technique has been applied successively to five proteins of mass 12.2 kDa to 18.3 kDa, yielding segments of continuous sequence as long as 41 residues without the need for prior proteolytic fragmentation. Intact crosslinks such as disulfides or heme linkages interrupt the generation of these data.

Locating and identifying posttranslational modifications by in-source decay during MALDI-TOF mass spectrometry.

A technique is described for identifying and locating posttranslational modifications (PTMs) in peptides and proteins of known sequence by interpretation of c(n) ion signals generated by in-source decay during delayed ion extraction in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Sites of phosphorylation in seven synthetic peptides were determined, as was the location of both the heme group and N,N,N-trimethyllysine in yeast cytochrome c. A semi-automated data analysis process facilitates the identification of segments of the sequence on each side of the PTM, permitting its placement at the junction of the segments and definition of the added mass. A graphical display facilitates illustration of both the location and mass of the PTM.

Topographic study of arrestin using differential chemical modifications and hydrogen/deuterium exchange.

Arrestin is involved in the quenching of phototransduction by binding to photoactivated and phosphorylated rhodopsin (P-Rho*). To study its conformational changes and regions interacting with P-Rho*, arrestin was subjected to (1) differential acetylation at lysine residues in the presence and absence of P-Rho*, and (2) amide hydrogen/deuterium exchange. Labeled protein was proteolysed and analyzed by mass spectrometry. Three Lys residues, 28, 176, and 211, were protected from acetylation in native arrestin, although they were not located in regions exhibiting slow amide hydrogen exchange rates. The presence of P-Rho* protected lysine 201 from acetylation and partially protected 14 other lysyl residues, including (2, 5), (163, 166, 167), (232, 235, 236, 238), (267, 276), (298, 300), and 367, where parentheses indicate lysine residues found within the same peptide. In contrast, in the C-terminal region of arrestin, lysyl residues (386, 392, 395) were more exposed upon binding to P-Rho*. These data allowed us to identify functional regions in the arrestin molecule.

Microheterogeneity of human filaggrin: analysis of a complex peptide mixture using mass spectrometry.

Filaggrin is the product of posttranslational processing of the large, epidermal protein profilaggrin, which consists of 10 or more tandem filaggrin domains plus an amino and a carboxyl domain. According to fragmentary cDNA sequences, the filaggrin domains in the human protein vary at 40% of the amino acid positions; hence, mature filaggrin is a population of homologous but heterogeneous proteins, even within one individual. Available gene sequences give only a limited picture of the heterogeneity of human filaggrin protein because no complete human profilaggrin gene has been sequenced. Questions about the extent of heterogeneity of filaggrin within and between individuals have not been answered, nor have questions concerning the limited proteolytic cleavage of human profilaggrin that generates filaggrin in vivo. In order to address these questions and to provide an analysis of the primary structure of human filaggrins, we employed various methods of mass spectrometry. The intact protein and a tryptic digest of the mixture of human filaggrins were examined by matrix-assisted laser desorption time-of-flight mass spectrometry. Tryptic digests of human filaggrin from single individuals were also separated and analyzed by liquid chromatography/mass spectrometry (LC/MS) (using electrospray mass spectrometry), and specific peptides were identified by tandem mass spectrometry (MS/MS). A robust data analysis program, Sherpa, was developed to facilitate the interpretation of both LC/MS and MS/MS. These experiments show that human filaggrin includes heterogeneity not yet seen in cDNA sequences, but that much structure is highly conserved. Interestingly, we found that the heterogeneity is conserved among individuals. An approximation of the regions linking filaggrins in human profilaggrin is developed. These investigations provide a unique test of the limits of tryptic mapping of complex mixtures using mass spectrometry.