Neutrophil Gelatinase-Associated Lipocalin Cutoff Value Selection and Acute Kidney Injury Classification System Determine Phenotype Allocation and Associated Outcomes.

Background: We explored the extent to which neutrophil gelatinase-associated lipocalin (NGAL) cutoff value selection and the acute kidney injury (AKI) classification system determine clinical AKI-phenotype allocation and associated outcomes. Methods: Cutoff values from ROC curves of data from two independent prospective cardiac surgery study cohorts (Magdeburg and Berlin, Germany) were used to predict Kidney Disease: Improving Global Outcome (KDIGO)- or Risk, Injury, Failure, Loss of kidney function, End-stage (RIFLE)-defined AKI. Statistical methodologies (maximum Youden index, lowest distance to [0, 1] in ROC space, sensitivity≍specificity) and cutoff values from two NGAL meta-analyses were evaluated. Associated risks of adverse outcomes (acute dialysis initiation and in-hospital mortality) were compared. Results: NGAL cutoff concentrations calculated from ROC curves to predict AKI varied according to the statistical methodology and AKI classification system (10.6-159.1 and 16.85-149.3 ng/mL in the Magdeburg and Berlin cohorts, respectively). Proportions of attributed subclinical AKI ranged 2%-33.0% and 10.1%-33.1% in the Magdeburg and Berlin cohorts, respectively. The difference in calculated risk for adverse outcomes (fraction of odds ratios for AKI-phenotype group differences) varied considerably when changing the cutoff concentration within the RIFLE or KDIGO classification (up to 18.33- and 16.11-times risk difference, respectively) and was even greater when comparing cutoff methodologies between RIFLE and KDIGO classifications (up to 25.7-times risk difference). Conclusions: NGAL positivity adds prognostic information regardless of RIFLE or KDIGO classification or cutoff selection methodology. The risk of adverse events depends on the methodology of cutoff selection and AKI classification system.

A Picture is Worth 1000 Words: Teaching Science Communication with Graphical Abstract Assignments.

An important part of scientific training in undergraduate curriculum involves teaching students how to effectively communicate in science. Scientific writing and oral presentations are important parts of most science classes. The same cannot be said about a new and emerging aspect of many recent scientific articles: graphical abstracts. In recent years, many scientific journals have adopted graphical abstracts as a way to capture both the scientific audience and increase visibility on social media platforms. Graphical abstracts are becoming the norm for many journals; however, there is no equivalent training in undergraduate classes that teaches students the intricate skills of efficient graphical design. In this paper, we share our course design and discuss how students can be taught to design better experiments and excel in communicating their research findings through graphical abstracts.

[Endoscopic Ultrasound Drainage of the Gallbladder in Acute Cholecystitis in Patients at High Surgical Risk]. / Endosonografische Drainage der Gallenblase wegen akuter Cholezystitis bei Patienten mit hohem Operationsrisiko.

BACKGROUND: The standard treatment for acute cholecystitis is laparoscopic cholecystectomy. Alternative procedures are used for patients at high surgical risk. Percutaneous drainage is widely available. The alternative of transpapillary drainage of the gallbladder via the ductus cysticus has only limited prospects of success. With the widespread use of interventional endoscopic ultrasound and the development of new stent systems, endoscopic ultrasound gallbladder drainage has proven to be a safe and reliable procedure. MATERIAL AND METHOD: We retrospectively report on our experiences in 11 consecutive patients with endoscopic ultrasound gallbladder drainage in acute cholecystitis between December 2018 and January 2021. RESULTS: 11 patients with acute cholecystitis with a mean age of 84.5 years (70-95 years) are reported. All patients had severe general comorbidities or advanced abdominal tumours or a combination of these conditions. After interdisciplinary debate, the indication for interventional therapy was made. This was carried out in 9 cases by means of endosonographic drainage alone and in 2 cases by means of percutaneous and two-stage endosonographic drainage. Technical success was achieved in 10 cases (91%), clinical success in 9 cases (82%). In 2 cases there were procedural complications that led to the operation. CONCLUSION: In the case of high surgical risks, endosonographic drainage of the gall bladder is a safe and definitive therapy. This can be performed alone or in combination with percutaneous drainage. Endoscopic ultrasound drainage is superior to percutaneous drainage alone, due to its lower complication rates and lower rates of necessary follow-up interventions. Therefore, in cases of relatively high surgical risk, endoscopic ultrasound drainage of the gall bladder should be preferred to percutaneous drainage, especially when definitive therapy is required.

Gains in Scientific Identity, Scientific Self-Efficacy, and Career Intent Distinguish Upper-Level CUREs from Traditional Experiences in the Classroom.

Attrition from the science, technology, engineering, and math (STEM) pipeline limits the number of graduates needed to meet STEM workforce demand and impedes efforts to diversify the workforce. Identifying factors that underlie academic success and STEM persistence is an important component to increasing the number of STEM graduates. The current study utilizes the social influence process indicators of the Tripartite Integration Model of Social Influence to investigate effects of course-based undergraduate research experience (CURE) participation and to predict career intent in a diverse population. CURE participants experienced significant gains in scientific self-efficacy, scientific identity, and career intent, while students in control courses did not. Between-groups analysis showed that scientific self-efficacy and scientific identity increased significantly more for CURE participants than for non-CURE participants. Regression analysis revealed that scientific identity was the only significant predictor of a student's career intent. This work underscores the central importance of prioritizing scientific identity in STEM curricula to improve throughput in the STEM pipeline and illustrates the usefulness of CUREs as viable interventions to positively influence factors that promote STEM career intent.

Efficacy of Tourniquet Use in Total Knee Arthroplasty: A Retrospective Cohort Review.

Background: The use of a tourniquet has become widely accepted as standard practice during total knee arthroplasty (TKA). There are conflicting outcomes in using a tourniquet during TKA. This brings to question the role a tourniquet has in TKA. Therefore, we conducted a retrospective cohort study to examine the effects of TKA with and without the use of a tourniquet. Methods: A total of 120 patients (n = 60 underwent TKA with tourniquet and n = 60 underwent TKA without tourniquet) were included in this study. Patient medical records were retrospectively reviewed for preoperative and postoperative data. The Gross formula, a validated formula for calculating blood loss, was used to calculate each patient's total blood loss. Statistical analysis was performed using independent t-tests, Mann-Whitney U tests, and/or chi-square tests. Significance was determined using an alpha level of P < .05. Results: There was no statistically significant difference (P = .49) in the amount of total blood loss between patients undergoing TKA with a tourniquet and those without (199.6 ± 92.2 mL vs 211.1 ± 88.1 mL, respectively). However, there were statistically significant differences in the operating room time (P = .005), surgery time (P = .008), and functional return of postoperative straight leg raise (P < .001) between groups. Conclusions: This study supports existing evidence that tourniquet use during TKA does not significantly alter blood loss and presents evidence that using a tourniquet during TKA may add additional cost and increase surgical time without benefit.

Failure of Screw/Shell Interface in the Trident II Acetabular System in Total Hip Arthroplasty.

We report a case series of 2 patients with screw/shell interface failure in the Stryker Trident II Acetabular System. Both failures consisted of screw penetration through the Trident II acetabular shell. One failure was observed postoperatively after a revision from a cephalomedullary nail to a total hip arthroplasty while the other was observed intraoperatively during a primary total hip arthroplasty. Both component failures were managed conservatively with weight-bearing as tolerated and radiographic monitoring. These are the first reported cup/screw failures of the Stryker Trident II system and should raise awareness of the potential complication and implant design flaw. When placing acetabular screws, we recommend obtaining intraoperative orthogonal screw radiographs that are tangential to the shell surface to assess for screw/shell failure.

Proteomic analysis of the acidocalcisome, an organelle conserved from bacteria to human cells.

Acidocalcisomes are acidic organelles present in a diverse range of organisms from bacteria to human cells. In this study acidocalcisomes were purified from the model organism Trypanosoma brucei, and their protein composition was determined by mass spectrometry. The results, along with those that we previously reported, show that acidocalcisomes are rich in pumps and transporters, involved in phosphate and cation homeostasis, and calcium signaling. We validated the acidocalcisome localization of seven new, putative, acidocalcisome proteins (phosphate transporter, vacuolar H+-ATPase subunits a and d, vacuolar iron transporter, zinc transporter, polyamine transporter, and acid phosphatase), confirmed the presence of six previously characterized acidocalcisome proteins, and validated the localization of five novel proteins to different subcellular compartments by expressing them fused to epitope tags in their endogenous loci or by immunofluorescence microscopy with specific antibodies. Knockdown of several newly identified acidocalcisome proteins by RNA interference (RNAi) revealed that they are essential for the survival of the parasites. These results provide a comprehensive insight into the unique composition of acidocalcisomes of T. brucei, an important eukaryotic pathogen, and direct evidence that acidocalcisomes are especially adapted for the accumulation of polyphosphate.

The acidocalcisome vacuolar transporter chaperone 4 catalyzes the synthesis of polyphosphate in insect-stages of Trypanosoma brucei and T. cruzi.

Polyphosphate is a polymer of inorganic phosphate found in both prokaryotes and eukaryotes. Polyphosphate typically accumulates in acidic, calcium-rich organelles known as acidocalcisomes, and recent research demonstrated that vacuolar transporter chaperone 4 catalyzes its synthesis in yeast. The human pathogens Trypanosoma brucei and T. cruzi possess vacuolar transporter chaperone 4 homologs. We demonstrate that T. cruzi vacuolar transporter chaperone 4 localizes to acidocalcisomes of epimastigotes by immunofluorescence and immuno-electron microscopy and that the recombinant catalytic region of the T. cruzi enzyme is a polyphosphate kinase. RNA interference of the T. brucei enzyme in procyclic form parasites reduced short chain polyphosphate levels and resulted in accumulation of pyrophosphate. These results suggest that this trypanosome enzyme is an important component of a polyphosphate synthase complex that utilizes ATP to synthesize and translocate polyphosphate to acidocalcisomes in insect stages of these parasites.

Trypanosoma brucei vacuolar transporter chaperone 4 (TbVtc4) is an acidocalcisome polyphosphate kinase required for in vivo infection.

Polyphosphate (polyP) is an anionic polymer of orthophosphate groups linked by high energy bonds that typically accumulates in acidic, calcium-rich organelles known as acidocalcisomes. PolyP synthesis in eukaryotes was unclear until it was demonstrated that the protein named Vtc4p (vacuolar transporter chaperone 4) is a long chain polyP kinase that localizes to the yeast vacuole. Here, we report that TbVtc4 (Vtc4 ortholog of Trypanosoma brucei) encodes, in contrast, a short chain polyP kinase that localizes to acidocalcisomes. The subcellular localization of TbVtc4 was demonstrated by fluorescence and electron microscopy of cell lines expressing TbVtc4 in its endogenous locus fused to an epitope tag and by purified polyclonal antibodies against TbVtc4. Recombinant TbVtc4 was expressed in bacteria, and polyP kinase activity was assayed in vitro. The in vitro growth of conditional knock-out bloodstream form trypanosomes (TbVtc4-KO) was significantly affected relative to the parental cell line. This mutant had reduced polyP kinase activity and short chain polyP content and was considerably less virulent in mice. The wild-type phenotype was recovered when an ectopic copy of the TbVtc4 gene was expressed in the presence of doxycycline. The mutant also exhibited a defect in volume recovery under osmotic stress conditions in vitro, underscoring the relevance of polyP in osmoregulation.

Identification of contractile vacuole proteins in Trypanosoma cruzi.

Contractile vacuole complexes are critical components of cell volume regulation and have been shown to have other functional roles in several free-living protists. However, very little is known about the functions of the contractile vacuole complex of the parasite Trypanosoma cruzi, the etiologic agent of Chagas disease, other than a role in osmoregulation. Identification of the protein composition of these organelles is important for understanding their physiological roles. We applied a combined proteomic and bioinfomatic approach to identify proteins localized to the contractile vacuole. Proteomic analysis of a T. cruzi fraction enriched for contractile vacuoles and analyzed by one-dimensional gel electrophoresis and LC-MS/MS resulted in the addition of 109 newly detected proteins to the group of expressed proteins of epimastigotes. We also identified different peptides that map to at least 39 members of the dispersed gene family 1 (DGF-1) providing evidence that many members of this family are simultaneously expressed in epimastigotes. Of the proteins present in the fraction we selected several homologues with known localizations in contractile vacuoles of other organisms and others that we expected to be present in these vacuoles on the basis of their potential roles. We determined the localization of each by expression as GFP-fusion proteins or with specific antibodies. Six of these putative proteins (Rab11, Rab32, AP180, ATPase subunit B, VAMP1, and phosphate transporter) predominantly localized to the vacuole bladder. TcSNARE2.1, TcSNARE2.2, and calmodulin localized to the spongiome. Calmodulin was also cytosolic. Our results demonstrate the utility of combining subcellular fractionation, proteomic analysis, and bioinformatic approaches for localization of organellar proteins that are difficult to detect with whole cell methodologies. The CV localization of the proteins investigated revealed potential novel roles of these organelles in phosphate metabolism and provided information on the potential participation of adaptor protein complexes in their biogenesis.

Target of rapamycin (TOR)-like 1 kinase is involved in the control of polyphosphate levels and acidocalcisome maintenance in Trypanosoma brucei.

Target of rapamycin (TOR) kinases are highly conserved protein kinases that integrate signals from nutrients and growth factors to coordinate cell growth and cell cycle progression. It has been previously described that two TOR kinases control cell growth in the protozoan parasite Trypanosoma brucei, the causative agent of African trypanosomiasis. Here we studied an unusual TOR-like protein named TbTOR-like 1 containing a PDZ domain and found exclusively in kinetoplastids. TbTOR-like 1 localizes to unique cytosolic granules. After hyperosmotic stress, the localization of the protein shifts to the cell periphery, different from other organelle markers. Ablation of TbTOR-like 1 causes a progressive inhibition of cell proliferation, producing parasites accumulating in the S/G(2) phase of the cell cycle. TbTOR-like 1 knocked down cells have an increased area occupied by acidic vacuoles, known as acidocalcisomes, and are enriched in polyphosphate and pyrophosphate. These results suggest that TbTOR-like 1 might be involved in the control of acidocalcisome and polyphosphate metabolism in T. brucei.

Calcium- and polyphosphate-containing acidocalcisomes in chicken egg yolk.

BACKGROUND INFORMATION: Poly P (inorganic polyphosphate) is a polymer formed by P(i) residues linked by high-energy phosphoanhydride bonds. The presence of poly P in bacteria, fungi, algae and protists has been widely recognized, but the distribution of poly P in more complex eukaryotes has been poorly studied. Poly P accumulates, together with calcium, in acidic vesicles or acidocalcisomes in a number of organisms and possesses a diverse array of functions, including roles in stress response, blood clotting, inflammation, calcification, cell proliferation and apoptosis. RESULTS: We report here that a considerable amount of phosphorus in the yolk of chicken eggs is in the form of poly P. DAPI (4',6-diamidino-2-phenylindole) staining showed that poly P is localized mainly in electron-dense vesicles located inside larger vacuoles (compound organelles) that are randomly distributed in the yolk. These internal vesicles were shown to contain calcium, potassium, sodium, magnesium, phosphorus, chlorine, iron and zinc, as detected by X-ray microanalysis and elemental mapping. These vesicles stain with the acidophilic dye Acridine Orange. The presence of poly P in organellar fractions of the egg yolk was evident in agarose gels stained with Toluidine Blue and DAPI. Of the total phosphate (Pi) of yolk organelles, 16% is present in the form of poly P. Total poly P content was not altered during the first 4 days of embryogenesis, but poly P chain length decreased after 1 day of development. CONCLUSIONS: The results of the present study identify a novel organelle in chicken egg yolk comprising acidic vesicles with a morphology, physiology and composition similar to those of acidocalcisomes, within larger acidic vacuoles. The elemental composition of these acidocalcisomes is proportionally similar to the elemental composition of the yolk, suggesting that most of these elements are located in these organelles, which might be an important storage compartment in eggs.

Evolution of acidocalcisomes and their role in polyphosphate storage and osmoregulation in eukaryotic microbes.

Acidocalcisomes are acidic electron-dense organelles, rich in polyphosphate (poly P) complexed with calcium and other cations. While its matrix contains enzymes related to poly P metabolism, the membrane of the acidocalcisomes has a number of pumps (Ca(2+)-ATPase, V-H(+)-ATPase, H(+)-PPase), exchangers (Na(+)/H(+), Ca(2+)/H(+)), and at least one channel (aquaporin). Acidocalcisomes are present in both prokaryotes and eukaryotes and are an important storage of cations and phosphorus. They also play an important role in osmoregulation and interact with the contractile vacuole complex in a number of eukaryotic microbes. Acidocalcisomes resemble lysosome-related organelles (LRO) from mammalian cells in many of their properties. They share similar morphological characteristics, acidic properties, phosphorus contents and a system for targeting of their membrane proteins through adaptor complex-3 (AP-3). Storage of phosphate and cations may represent the ancestral physiological function of acidocalcisomes, with cation and pH homeostasis and osmoregulatory functions derived following the divergence of prokaryotes and eukaryotes.

Contrast enhanced sonography shows superior microvascular renal allograft perfusion in patients switched from cyclosporine A to everolimus.

BACKGROUND: Real-time contrast enhanced sonography (CES) provides quantitative information on microvascular tissue perfusion in renal allografts. In contrast to calcineurin inhibitors, mammalian target of rapamycin inhibitors may have beneficial effects on renal microvascular tissue perfusion. There is no information on the microperfusion of renal allografts in patients receiving either mammalian target of rapamycin inhibitor or calcineurin inhibitor. METHODS: In a prospective randomized, clinical trial, renal parenchymal tissue perfusion of 24 stable renal allograft recipients was evaluated with CES. Eleven patients were kept on cyclosporine A (CsA); 13 were converted to everolimus (EVR). Measurements were made at the time of the switch from CsA to EVR, 8.21+/-6.36 months posttransplantation, and 21.2+/-6.57 months posttransplantation. In addition to laboratory and clinical parameters, Doppler indices and estimated glomerular filtration rate (eGFR) were measured. RESULTS.: After the switch from CsA to EVR, microvascular perfusion in the EVR-treated patients (Axbeta value at baseline 9.23+/-7.44 dB/sec, Axbeta value at time of follow-up 19.6+/-13.0 dB/sec, P=0.03) and the estimated GFR (81.2+/-20.3 and 96.9+/-22.6 mL/min, P=0.001) improved significantly. Microvascular perfusion (Axbeta 7.04+/-5.32 dB/sec and Axbeta 8.66+/-9.01 dB/sec, P=0.34) and the eGFR of the group continuing CsA treatment remained stable (78.5+/-25.9 and 73.2+/-37.3 mL/min, P=0.1). CONCLUSION: The study demonstrates that renal microperfusion visualized by CES based on microbubble contrast agent and concomitantly kidney function, improved significantly after the switch from CsA to EVR.

Low levels of prothrombin time (INR) and platelets do not increase the risk of significant bleeding when placing central venous catheters.

BACKGROUND AND PURPOSE: Central venous catheters are frequently placed in intensive care medicine for multiple indications. The risk of severe bleeding after cannulation is considered to be increased in patients with abnormal coagulation, common in critically ill patients. PATIENTS AND METHODS: This open prospective trial, performed at two medical intensive care units and one hematology intermediate care ward, investigated whether insertion of a central venous catheter in patients with coagulopathy (prothrombin time or= 1.5] and/or platelets