Update of Contrast-enhanced Ultrasound in Musculoskeletal Medicine: Clinical Perspectives - A Review.

Contrast-enhanced ultrasound (CEUS) uses an intravascular contrast agent to enhance blood flow signals and assess microcirculation in different parts of the human body. Over the past decade, CEUS has become more widely applied in musculoskeletal (MSK) medicine, and the current review aims to systematically summarize current research on the application of CEUS in the MSK field, focusing on 67 articles published between January 2001 and June 2021 in online databases including PubMed, Scopus, and Embase. CEUS has been widely used for the clinical assessment of muscle microcirculation, tendinopathy, fracture nonunions, sports-related injuries, arthritis, peripheral nerves, and tumors, and can serve as an objective and quantitative evaluation tool for prognosis and outcome prediction. Optimal CEUS parameters and diagnostic cut off values for each disease category remain to be confirmed.

Platelet dysfunction persists after trauma despite balanced blood product resuscitation.

BACKGROUND: Platelet activation and aggregation are critical to the initiation of hemostasis after trauma with hemorrhage. Platelet dysfunction is a well-recognized phenomenon contributing to trauma-induced coagulopathy. The goal of this study was to evaluate the timing and severity of platelet dysfunction in massively transfused, traumatically injured patients during the first 72 hours after injury and its association with 30-day survival. METHODS: A retrospective secondary cohort study of platelet count and function was performed using samples from the Pragmatic Randomized Optimal Platelet and Plasma Ratios trial. Platelet characteristics were measured at 8 timepoints during the first 72 hours of hospitalization and compared between 30-day survivors and nonsurvivors. Platelet counts were assessed via flow cytometry. Platelet function was analyzed with the use of serial thrombelastography and impedance aggregometry with agonists arachidonic acid, adenosine diphosphate, collagen, thrombin receptor activating peptide, and ristocetin. RESULTS: In total, 680 patients were included for analysis. Platelet counts were significantly lower from baseline to 72 hours after hospital admission with further 1.3 to 2-fold reductions noted in nonsurvivors compared to survivor patients. Platelet aggregation via adenosine diphosphate, arachidonic acid, collagen, thrombin receptor activating peptide, and ristocetin was significantly lower in nonsurvivors at all time points. The nadir of platelet aggregation was 2 to 6 hours after admission with significant improvements in viscoelastic maximum clot formation and agonist-induced aggregation by 12 hours without concomitant improvement in platelet count. CONCLUSION: Platelet aggregability recovers 12 hours after injury independent of worsening thrombocytopenia. Failure of platelet function to recover portends a poor prognosis.

Granulocyte-macrophage colony stimulating factor enhances efficacy of nimustine rendezvousing with temozolomide plus irradiation in patients with glioblastoma.

BACKGROUND: Glioblastoma is the most common and most aggressive type of primary brain tumor. OBJECTIVE: The aim of this study was to investigate the efficacy and safety of intranasal granulocyte-macrophage colony stimulating factor (GM-CSF) administration combined with chemoradiotherapy in patients with glioblastoma who underwent surgery. METHODS: Ninety-two patients were randomly divided into two groups: a control group (n= 46), who received radiotherapy with adjuvant local delivery of nimustine hydrochloride (ACNU) and systemic administration of temozolomide, and an intervention group (n= 46), who received intranasal GM-CSF prior to each cycle of adjuvant chemotherapy in addition to the treatment of the control group. Karnofsky performance status (KPS) scores, progression-free survival (PFS), overall survival (OS), and adverse effects were calculated and compared between the two groups. RESULTS: Compared with the control group, the intervention group had longer PFS (7.8 vs. 6.9 months, P= 0.016) and OS (19.2 vs. 17.1 months, P= 0.045, without adjustment for interim analyses). The KPS scores were also higher in the intervention group than in the control group after 6 months (84.35 ± 8.86 vs. 80.65 ± 7.72; t= 4.552, P= 0.036). Furthermore, the patients in the intervention group had lower incidence of neutropenia and thrombocytopenia (8.7% vs. 29.5%, P= 0.012; 8.7% vs. 18.2%, P= 0.186). Other adverse events were similar in both groups, and most adverse events were grade I/II and resolved spontaneously. CONCLUSION: Intranasal GM-CSF enhances the efficacy of the local ACNU administration combined with oral temozolomide chemotherapy. The survival and performance status were significantly improved in patients with glioblastoma after surgery. Additionally, the GM-CSF therapy was able to reduce the occurrence of chemotherapy-related neutropenia and thrombocytopenia.

Association of Changes in Antithrombin Activity Over Time With Responsiveness to Enoxaparin Prophylaxis and Risk of Trauma-Related Venous Thromboembolism.

Importance: Venous thromboembolism (VTE) affects 2% to 20% of recovering trauma patients, despite aggressive prophylaxis with enoxaparin. Antithrombin is a primary circulating anticoagulant and crucial component of enoxaparin thromboprophylaxis. Approximately 20% of trauma patients present with antithrombin deficiency (antithrombin activity <80%). Objective: To examine time-dependent changes in antithrombin activity, responsiveness to enoxaparin, as measured by anti-factor Xa (anti-FXa) levels, and incidence of VTE after severe trauma and to assess the association of ex vivo antithrombin supplementation with patients' sensitivity to enoxaparin prophylaxis. Design, Setting, and Participants: This single-center, prospective cohort study was performed at a level 1 trauma center between January 7, 2019, and February 28, 2020. Adult trauma patients admitted to the trauma service at high risk for VTE, based on injury pattern and severity, were screened and enrolled. Patients who were older than 70 years, were pregnant, had a known immunologic or coagulation disorder, or were receiving prehospital anticoagulants were excluded. Exposures: Blood samples were collected on emergency department arrival and daily for the first 8 days of hospitalization. Main Outcomes and Measures: Patients' antithrombin activity and anti-FXa levels were measured by a coagulation analyzer, and thrombin generation was measured by calibrated automated thrombography. Responsiveness to enoxaparin was assessed by measuring anti-FXa levels 4 to 6 hours after the first daily enoxaparin dose and compared between patients who developed VTE and who did not. In addition, the associations of ex vivo supplementation of antithrombin with plasma anti-FXa levels were assessed. Results: Among 150 patients enrolled (median [IQR] age, 35 [27-53] years; 37 [24.7%] female and 113 [75.3%] male; 5 [3.3%] Asian, 32 [21.3%] Black, and 113 [75.3%] White; and 51 [34.0%] of Hispanic ethnicity), 28 (18.7%) developed VTE. Patients with VTE had significantly lower antithrombin activity on admission compared with patients without VTE (median [IQR], 91% [79%-104%] vs 100% [88%-112%]; P = .04), as well as lower antithrombin activity on hospital days 5 (median (IQR), 90% [83%-99%] vs 114% [99%-130%]; P = .011), 6 (median [IQR], 97% [81%-109%] vs 123% [104%-134%]; P = .003), 7 (median [IQR], 82% [74%-89%] vs 123% [110%-140%]; P < .001), and 8 (median [IQR], 99% [85%-100%] vs 123% [109%-146%]; P = .011). Anti-FXa levels were significantly lower in patients with VTE vs those without VTE at hospital day 4 (median [IQR], 0.10 [0.05-0.14] IU/mL vs 0.18 [0.13-0.23] IU/mL; P = .006), day 6 (median [IQR], 0.12 [0.08-0.14] IU/mL vs 0.22 [0.13-0.28] IU/mL; P = .02), and day 7 (median [IQR], 0.11 [0.08-0.12] IU/mL vs 0.21 [0.13, 0.28] IU/mL; P = .002). Multivariable analyses found that for every 10% decrease in antithrombin activity during the first 3 days, the risk of VTE increased 1.5-fold. Conclusions and Relevance: The results of this cohort study suggest that after severe trauma, antithrombin deficiency is common and contributes to enoxaparin resistance and VTE. Interventional studies are necessary to determine the efficacy of antithrombin supplementation in the reduction of VTE incidence.

Survival analysis by inflammatory biomarkers in severely injured patients undergoing damage control resuscitation.

BACKGROUND: Although early balanced blood product resuscitation has improved mortality after traumatic injury, many patients still suffer from inflammatory complications. The goal of this study was to identify inflammatory mediators associated with death and multiorgan system failure following severe injury after patients undergo blood product resuscitation. METHODS: A retrospective secondary analysis of inflammatory markers from the Pragmatic Randomized Optimal Platelet and Plasma Ratios study was performed. Twenty-seven serum biomarkers were measured at 8 time points in the first 72 hours of care and were compared between survivors and nonsurvivors. Biomarkers with significant differences were further analyzed by adjudicated cause of 30-day mortality. RESULTS: Biomarkers from 680 patients were analyzed. Seven key inflammatory markers (IL-1ra, IL-6, IL-8, IL-10, eotaxin, IP-10, and MCP-1) were further analyzed. These cytokines were also noted to have the highest hazard ratios of death. Stepwise selection was used for multivariate analysis of survival by time point. MCP-1 at 2 hours, eotaxin and IP-10 at 12 hours, eotaxin at 24 hours, and IP-10 at 72 hours were associated with all-cause mortality. CONCLUSION: Early systemic inflammatory markers are associated with increased risk of mortality after traumatic injury. Future studies should use these biomarkers to prospectively calculate risks of morbidity and causes of mortality for all trauma patients.

The Impact of Rapid Infuser Use on the Platelet Count, Platelet Function, and Hemostatic Potential of Whole Blood.

BACKGROUND: Rapid infusion pumps employing filters, roller pumps, and heat exchangers for the administration of blood products are not approved for platelets or cryoprecipitate. This technology may decrease platelet count and degrade coagulation proteins. The effect of rapid infusers on the hemostatic potential of whole blood is unknown. METHODS: Five units of low titer O+ whole blood were obtained from anonymous donors. Each unit was subjected to infusion by five different techniques: (1) gravity infusion without a filter, (2) gravity infusion with a filter, (3) Belmont rapid infuser at 70 mL/min, (4) Belmont at 100 mL/min, and (5) pressurized infusion with a pneumatic pressure bag and filter. After infusion, platelet count, platelet function, thrombin generation, and hemostatic potential were measured for each aliquot. Infusion techniques were compared, using gravity infusion without a filter as the control. RESULTS: There was a significant decrease in platelet count from baseline (168,000) in the BELMONT70 (97,000) and BELMONT100 (94,000) groups (P < 0.05). However, there were no differences in platelet function (all P > 0.20). While there were no differences in thromboelastography parameters between control and infusion models (all P > 0.20), there were significant increases in thrombin generation parameters by CAT in both the BELMONT70 and BELMONT100 groups (all P < 0.05). CONCLUSIONS: The use of a rapid infuser decreases the platelet count of WB but does not decrease platelet function or overall hemostatic potential. In fact, thrombin generation and thrombin potential are actually increased. Rapid infusers are safe for the transfusion of WB.

Urinary cell cycle arrest proteins urinary tissue inhibitor of metalloprotease 2 and insulin-like growth factor binding protein 7 predict acute kidney injury after severe trauma: A prospective observational study.

BACKGROUND: Recognition and clinical diagnosis of acute kidney injury (AKI) after trauma is difficult. The majority of trauma patients do not have a known true baseline creatinine, which makes application of the guidelines set forth by the international guidelines difficult to apply. Use of alternative biomarkers of renal dysfunction in trauma patients may be beneficial. We hypothesized that urinary tissue inhibitor of metalloprotease 2 (TIMP-2) × insulin-like growth factor binding protein 7 (IGFBP-7) would accurately predict AKI development in severely injured trauma patients. METHODS: A prospective observational study of adult (≥16 years old) trauma intensive care unit (ICU) patients was performed between September 2018 to March 2019. Urine was collected on ICU admission and was measured for TIMP-2 × IGFBP-7. Univariate, multivariable, and receiver operating characteristic curve analyses were performed using the optimal threshold generated by a Youden index. MAIN RESULTS: Of 88 included patients, 75% were male, with a median injury severity score was 27 (interquartile range [IQR], 17-34), and age of 40 years (IQR, 28-54 years). Early AKI developed in 39 patients (44%), and of those, 7 (8%) required dialysis within 48 hours. Patients without early AKI had a TIMP-2 × IGFBP-7 of 0.17 U (IQR, 0.1-0.3 U), while patients with early AKI had a TIMP-2 × IGFBP-7 of 0.46 U (IQR, 0.17-1.29 U; p < 0.001). On multivariable analyses, TIMP-2 × IGFBP-7 was associated with AKI development (p = 0.02) and need for dialysis (p = 0.03). Using the optimal threshold 0.33 U to predict AKI, the area under the receiver operating characteristic curve was 0.731, with an accuracy of 0.75, sensitivity of 0.72, and specificity of 0.78. CONCLUSION: Urinary TIMP-2 × IGFBP-7 measured on ICU admission accurately predicted 48-hour AKI and was independently associated with AKI and dialysis requirement after trauma and is a promising screening tool for treatment. LEVEL OF EVIDENCE: Prognostic, prospective, observational study, level III.

Hemostatic potential of cold-stored non-leukoreduced whole blood over time: An assessment of platelet function and thrombin generation for optimal shelf life.

OBJECTIVES: Cold-stored low-titer whole blood (WB) is becoming increasingly used as the preferred product for initial hemorrhagic shock resuscitation. The purpose of this study was to identify whether the current 21-day shelf life is the optimal duration for storage of WB, maintaining hemostatic efficacy. METHODS: Five units of fresh low-titer group O WB (non-leukoreduced) were acquired from our regional blood center. These units were stored at 4°C for up to 21 days as per current clinical storage guidelines in our emergency department. Hemostatic parameters were measured in vitro at 0 days, 7 days, 14 days, and 21 days. Assessments of hemostatic potential included cell count, rapid thrombelastography (r-TEG) and kaolin thrombelastography (TEG), multiplate impedance aggregometry, and calibrated automated thrombogram (CAT). Univariate analysis, including one-way analysis of variance with repeated measures, was performed (STATA 12.1). RESULTS: Compared with baseline product (0 days), both platelet count and platelet function of WB showed sharp decreases at 7 days and again at 14 days. Platelet function deterioration was noted by r-TEG c (MA), TEG-MA, and multiplate arachidonic acid and adenosine diphosphate (all p < 0.001). With respect to clot initiation, r-TEG ACT and TEG R-time were similar over the 21-day shelf life (p = 0.058 and p = 0.620, respectively). Thrombin generation assessed by CAT demonstrated stable endogenous thrombin potential over the course of storage (p = 0.162), but increased peak thrombin generation and quicker time to peak generation after 7 days. CONCLUSION: While the platelet function of WB degrades significantly at 7 days (and again at 14 days), clot initiation remains stable over time, and thrombin generation appears to be improved at 7 days. This study supports a current storage limit for cold-stored, low-titer WB of 14 days.

Clinical Cellular Therapeutics Accelerate Clot Formation.

Clinical cellular therapeutics (CCTs) have shown preliminary efficacy in reducing inflammation after trauma, preserving cardiac function after myocardial infarction, and improving functional recovery after stroke. However, most clinically available cell lines express tissue factor (TF) which stimulates coagulation. We sought to define the degree of procoagulant activity of CCTs as related to TF expression. CCT samples from bone marrow, adipose, amniotic fluid, umbilical cord, multi-potent adult progenitor cell donors, and bone marrow mononuclear cells were tested. TF expression and phenotype were quantified using flow cytometry. Procoagulant activity of the CCTs was measured in vitro with thromboelastography and calibrated thrombogram. Fluorescence-activated cell sorting (FACS) separated samples into high- and low-TF expressing populations to isolate the contribution of TF to coagulation. A TF neutralizing antibody was incubated with samples to demonstrate loss of procoagulant function. All CCTs tested expressed procoagulant activity that correlated with expression of tissue factor. Time to clot and thrombin formation decreased with increasing TF expression. High-TF expressing cells decreased clotting time more than low-TF expressing cells when isolated from a single donor using FACS. A TF neutralizing antibody restored clotting time to control values in some, but not all, CCT samples. CCTs demonstrate wide variability in procoagulant activity related to TF expression. Time to clot and thrombin formation decreases as TF load increases and this procoagulant effect is neutralized by a TF blocking antibody. Clinical trials using CCTs are in progress and TF expression may emerge as a safety release criterion. Stem Cells Translational Medicine 2018;7:731-739.

Enhanced antitumor effects of radiotherapy combined local nimustine delivery rendezvousing with oral temozolomide chemotherapy in glioblastoma patients.

BACKGROUND: Glioblastoma (GBM) is one of the worst cancers with bad prognosis despite systemic chemotherapy and radiotherapy after surgery. METHODS: In this study, 71 patients with GBM were enrolled and randomly assigned to two groups: Receiving radiotherapy with concomitant and adjuvant temozolomide (TMZ) (TMZ, standard therapy) after surgery, or receiving radiotherapy with concomitant and adjuvant local delivery of nimustine (ACNU) rendezvousing with oral TMZ (rendezvous therapy). In the follow-up of all patients and the progression-free survival (PFS), overall survival (OS), Karnofsky performance score (KPS) and toxicities were recorded. RESULTS: For the whole cohort, the median OS was 18.0 months, and the median PFS was 7.8 months. A significantly longer OS was observed in patients received rendezvous therapy than those who receiving standard therapy (18.5 months vs. 16.0 months; P = 0.014), as well as PFS (8.8 months vs. 7.0 months; P = 0.008). The KPS ≥70 rates were 81.8%, 40.9%, 20.5% in 1, 2, and 3 years for the rendezvous therapy group, significantly superior to standard therapy group. The most common toxicities were tolerable gastrointestinal reaction, liver dysfunction, and hematological toxicities, which were relieved with symptomatic treatment. Grade 3 or 4 toxicity was documented in 8 (18.3%) patients in rendezvous therapy group, while it was observed in 6 (22.2%) patients in standard therapy group during whole treatment process. CONCLUSIONS: Compared to standard therapy, the antitumor effects of rendezvous therapy were more effective in GBM patients without increasing the toxicities.

Never-frozen liquid plasma blocks endothelial permeability as effectively as thawed fresh frozen plasma.

BACKGROUND: Thawed fresh frozen plasma (TP) is a preferred plasma product for resuscitation but can only be used for up to 5 days after thawing. Never-frozen, liquid plasma (LQP) is approved for up to 26 days when stored at 1°C to 6°C. We have previously shown that TP repairs tumor necrosis factor α (TNF-α)-induced permeability in human endothelial cells (ECs). We hypothesized that stored LQP repairs permeability as effectively as TP. METHODS: Three single-donor LQP units were pooled. Aliquots were frozen, and samples were thawed on Day 0 (TP0) then refrigerated for 5 days (TP5). The remaining LQP was kept refrigerated for 28 days, and aliquots were analyzed every 7 days. The EC monolayer was stimulated with TNF-α (10 ng/mL), inducing permeability, followed by a treatment with TP0, TP5, or LQP aged 0, 7, 14, 21, and 28 days. Permeability was measured by leakage of fluorescein isothiocyanate-dextran through the EC monolayer. Hemostatic profiles of samples were evaluated by thrombogram and thromboelastogram. Statistical analysis was performed using two-way analysis of variance, with p < 0.05 deemed significant. RESULTS: TNF-α increased permeability of the EC monolayer twofold compared with medium control. There was a significant decrease in permeability at 0, 7, 14, 21, and 28 days when LQP was used to treat TNF-α-induced EC monolayers (p < 0.001). LQP was as effective as TP0 and TP5 at reducing permeability. Stored LQP retained the capacity to generate thrombin and form a clot. CONCLUSION: LQP corrected TNF-α-induced EC permeability and preserved hemostatic potential after 28 days of storage, similar to TP stored for 5 days. The significant logistical benefit (fivefold) of prolonged LQP storage improves the immediate availability of plasma as a primary resuscitative fluid for bleeding patients.

Better hemostatic profiles of never-frozen liquid plasma compared with thawed fresh frozen plasma.

BACKGROUND: Immediate use of thawed fresh frozen plasma (FFP) when resuscitating hemorrhagic shock patients has become more common. According to the AABB (formerly known as American Association of Blood Banks), FFP is the preferred product that can be used up to 5 days after thawing. However, limited data exist on the clinical use and hemostatic profiles of Food and Drug Administration-approved liquid plasma (LQP), which can be stored at 1 °C to 6 °C for up to 26 days. We characterized changes in LQP hemostatic potential during 26 days of cold storage. METHODS: Ten FFP and 10 LQP single-donor units, matched by sex and blood group, were analyzed. FFP was thawed and kept refrigerated for 5 days and LQP for 26 days. Plasma samples were evaluated at Days 0 and 5 for thawed plasma (TP) and 0, 5, 10, 20, and 26 for LQP, by thrombelastography, thrombogram, platelet counts, platelet microparticles, clotting factors, and natural coagulation inhibitors. RESULTS: LQP had a better capacity to form a clot and generate thrombin compared with TP. LQP's hemostatic potential, expressed as endogenous thrombin potential (total amount of thrombin [nM] formed over time [minute]), initially exceeded that of TP (1,425 vs. 1,184, p < 0.05) but decreased to levels similar to TP by Day 26 (1,201 vs. 1,103, p = 0.15). Significantly higher platelet microparticles were found in LQP on Day 26 compared with those in LQP on Day 0 (23.6 x 10(9)/L vs. 3 x 10(9)/L, p < 0.001) or those in TP on Day 5 (2.8 x 10(9)/L). By Day 26, the majority of clotting factors and inhibitors retained more than 88% of their initial activities in LQP, with the few exceptions of factors well known to be unstable. CONCLUSION: The hemostatic profiles of LQP were better and sustained five times longer than the more commonly used TP, indicating that never-frozen plasma can be considered for use in the United States in trauma patients requiring immediate plasma resuscitation.