POLθ prevents MRE11-NBS1-CtIP-dependent fork breakage in the absence of BRCA2/RAD51 by filling lagging-strand gaps.

POLθ promotes repair of DNA double-strand breaks (DSBs) resulting from collapsed forks in homologous recombination (HR) defective tumors. Inactivation of POLθ results in synthetic lethality with the loss of HR genes BRCA1/2, which induces under-replicated DNA accumulation. However, it is unclear whether POLθ-dependent DNA replication prevents HR-deficiency-associated lethality. Here, we isolated Xenopus laevis POLθ and showed that it processes stalled Okazaki fragments, directly visualized by electron microscopy, thereby suppressing ssDNA gaps accumulating on lagging strands in the absence of RAD51 and preventing fork reversal. Inhibition of POLθ DNA polymerase activity leaves fork gaps unprotected, enabling their cleavage by the MRE11-NBS1-CtIP endonuclease, which produces broken forks with asymmetric single-ended DSBs, hampering BRCA2-defective cell survival. These results reveal a POLθ-dependent genome protection function preventing stalled forks rupture and highlight possible resistance mechanisms to POLθ inhibitors.

Stable kinetochore-microtubule attachment requires loop-dependent Ndc80-Ndc80 binding.

During cell division, kinetochores link chromosomes to spindle microtubules. The Ndc80 complex, a crucial microtubule binder, populates each kinetochore with dozens of copies. Whether adjacent Ndc80 complexes cooperate to promote microtubule binding remains unclear. Here we demonstrate that the Ndc80 loop, a short sequence that interrupts the Ndc80 coiled-coil at a conserved position, folds into a more rigid structure than previously assumed and promotes direct interactions between full-length Ndc80 complexes on microtubules. Mutations in the loop impair these Ndc80-Ndc80 interactions, prevent the formation of force-resistant kinetochore-microtubule attachments, and cause cells to arrest in mitosis for hours. This arrest is not due to an inability to recruit the kinetochore-microtubule stabilizing SKA complex and cannot be overridden by mutations in the Ndc80 tail that strengthen microtubule attachment. Thus, loop-mediated organization of adjacent Ndc80 complexes is crucial for stable end-on kinetochore-microtubule attachment and spindle assembly checkpoint satisfaction.

Smarcal1-Mediated Fork Reversal Triggers Mre11-Dependent Degradation of Nascent DNA in the Absence of Brca2 and Stable Rad51 Nucleofilaments.

Brca2 deficiency causes Mre11-dependent degradation of nascent DNA at stalled forks, leading to cell lethality. To understand the molecular mechanisms underlying this process, we isolated Xenopus laevis Brca2. We demonstrated that Brca2 protein prevents single-stranded DNA gap accumulation at replication fork junctions and behind them by promoting Rad51 binding to replicating DNA. Without Brca2, forks with persistent gaps are converted by Smarcal1 into reversed forks, triggering extensive Mre11-dependent nascent DNA degradation. Stable Rad51 nucleofilaments, but not RPA or Rad51T131P mutant proteins, directly prevent Mre11-dependent DNA degradation. Mre11 inhibition instead promotes reversed fork accumulation in the absence of Brca2. Rad51 directly interacts with the Pol α N-terminal domain, promoting Pol α and δ binding to stalled replication forks. This interaction likely promotes replication fork restart and gap avoidance. These results indicate that Brca2 and Rad51 prevent formation of abnormal DNA replication intermediates, whose processing by Smarcal1 and Mre11 predisposes to genome instability.

Modular assembly of RWD domains on the Mis12 complex underlies outer kinetochore organization.

Faithful chromosome segregation is mandatory for cell and organismal viability. Kinetochores, large protein assemblies embedded in centromeric chromatin, establish a mechanical link between chromosomes and spindle microtubules. The KMN network, a conserved 10-subunit kinetochore complex, harbors the microtubule-binding interface. RWD domains in the KMN subunits Spc24 and Spc25 mediate kinetochore targeting of the microtubule-binding subunits by interacting with the Mis12 complex, a KMN subcomplex that tethers directly onto the underlying chromatin layer. Here, we show that Knl1, a KMN subunit involved in mitotic checkpoint signaling, also contains RWD domains that bind the Mis12 complex and that mediate kinetochore targeting of Knl1. By reporting the first 3D electron microscopy structure of the KMN network, we provide a comprehensive framework to interpret how interactions of RWD-containing proteins with the Mis12 complex shape KMN network topology. Our observations unveil a regular pattern in the construction of the outer kinetochore.

The challenge of infectious diseases consultations in the emergency department: an Italian nationwide survey.

Diagnosis and management of infectious diseases (ID) at the emergency department (ED) are challenging due to the peculiar setting and the available diagnostic tools. The involvement of an ID consultant has been described to improve clinical outcomes and antimicrobial stewardship (AMS) programs. An online survey was sent to 100 Italian Departments of Infectious Diseases affiliated with the Italian Society of Infectious Diseases and Tropical Medicine (SIMIT). The primary objective of our study was to describe the characteristics of ID services in Italian EDs to identify possible challenges and shortcomings and provide tips to improve the management of patients. Secondary objectives included the evaluation of diagnostic capability and the management of patients with suspected or confirmed ID. Seventy-six out of the 100 SIMIT centers, 32 (42.1%) of which were teaching hospitals, answered the survey. In 62 (82.7%) centers, consultations were performed by the IDs specialist on call. In 29 (38.2%) centers, there was a formal AMS program, and 32 (42.7%) had protocols for antibiotic use in the ED. Microbiological tests to be performed before starting antibiotic treatment in the ED were clearly defined in 44 (57.9%) hospitals. This survey highlighted several challenges in the current organization of ID consultations in Italian EDs.

CD4/CD8 ratio in pregnant women with HIV and its association with pregnancy outcome: data from a national study in Italy.

PURPOSE: To evaluate associations between CD4/CD8 ratio and pregnancy outcomes in women with HIV. METHODS: We evaluated, in a national study of pregnant women with HIV receiving antiretroviral treatment (ART), values of CD4/CD8 ratio at entry in pregnancy, changes between first and third trimester, and possible associations with preterm delivery, low birthweight, and HIV-RNA < 50 copies/ml at third trimester in univariate and multivariate analyses. RESULTS: Among 934 women, 536 (57.4%) were already on ART at conception. CD4/CD8 ratio (baseline value 0.570) increased significantly between the first and third trimesters, particularly in women who started ART in pregnancy (+ 0.163, vs. + 0.036 in women already on treatment). The rate of CD4/CD8 ratio normalization, defined by achieving a ratio ≥ 1 at the third trimester, was 13.2%. In multivariable analyses, women who entered pregnancy with a CD4/CD8 ratio < 0.3, compared to women with ratio ≥ 1, were almost four-times less likely to have third-trimester HIV-RNA < 50 copies/ml (AOR 0.258, 95%CI 0.111-0.601), and more than twice as likely to have preterm delivery (AOR 2.379, 95%CI 1.082-5.232). For preterm delivery, also a baseline CD4/CD8 ratio between 0.3 and 0.45 was significantly associated with an increased risk (AOR: 3.415, 95%CI 1.690-6.900). CONCLUSION: We described for the first time independent associations of low CD4/CD8 ratio with preterm delivery and HIV-RNA suppression.

Pregnancy and neonatal outcomes in women with HIV-1 exposed to integrase inhibitors, protease inhibitors and non-nucleoside reverse transcriptase inhibitors: an observational study.

PURPOSE: Recommended regimens for pregnant women with HIV-1 are composed of two nucleoside reverse transcriptase inhibitors (NRTI) plus either a ritonavir-boosted protease inhibitor (PI) or an integrase strand transfer inhibitor (ISTI), with non-nucleoside reverse transcriptase inhibitors (NNRTI) representing an alternative drug class. The study's purpose was to compare these three options in terms of pregnancy outcomes. METHODS: Data from a national observational study of pregnant women with HIV-1 were used. The analysis included all pregnancies reported between 2008 and 2018, ending in live births and exposed within 32 weeks of gestation to three-drug regimens composed of a NRTI backbone plus a PI, a NNRTI or a ISTI, without class switching during pregnancy. Clinical and laboratory outcomes were evaluated in univariate and multivariable analyses. RESULTS: Overall, 794 exposed pregnancies were analyzed (PI 78.4%, NNRTI 15.4%, ISTI 6.2%). Almost all outcomes had similar rates in the three groups. Women who received PI in pregnancy were less likely to be virologically suppressed at third trimester. PI use was associated with higher bilirubin and triglyceride levels, and ISTI use with a lower rate of low birthweight. The differences in viral suppression at third trimester and in low birthweight were not maintained in multivariable analyses that were adjusted for confounders. DISCUSSION: We found no major differences in a wide range of outcomes relevant for pregnant women with HIV. Such results are reassuring, and this information may be helpful in a context of preconception counseling when therapeutic choices for pregnancy are discussed between women and care providers.

HIV-1 DNA dynamics and variations in HIV-1 DNA protease and reverse transcriptase sequences in multidrug-resistant patients during successful raltegravir-based therapy.

There is limited information on the variations of HIV-1 DNA mutation profile in reverse transcriptase (RT) and protease (PR) genes during suppressive antiretroviral treatment (plasma HIV-1 RNA continuously <50 copies/ml) with raltegravir (RAL)-based regimens in patients with baseline RT/PR resistant HIV. Twelve multidrug resistant (RT: 12/12, PR: 8/12) HIV-infected patients were followed during effectively suppressive RAL-based therapy. Total and integrated HIV-1 DNA were assessed by real time PCR at baseline and every 6 months. Ultrasensitive (threshold: 2.5 copies/ml) plasma HIV-1 RNA and genotypic analysis of RT and PR in proviral DNA were performed at baseline and at 24 months. Half of the patients had full viral suppression (plasma HIV-RNA < 2.5 copies/ml) at month 12. Total HIV-1 DNA declined significantly after 12 months of therapy (from 249.2 to 145.7 copies/106 cells, P = 0.023), and remained stable until 24 months, when total HIV-1 DNA levels raised, concomitantly with a less stringent suppression of HIV-1 RNA (81.8% of patients with >2.5 copies/ml). Integrated HIV-1 DNA did not show fluctuations during the study period. Sequencing of the PR and RT regions from HIV-1 DNA revealed changes in the resistance mutation profile in five patients. Total HIV-1 DNA declined after the introduction of RAL-based therapy, with a rebound after 2 years. No changes were observed in levels of integrated DNA, suggesting limited effect on archived HIV. The RT and PR sequence changes in archived HIV-1 DNA suggest that variation of the mutation profile can occur even in the absence of detectable HIV-1 RNA. J. Med. Virol. 88:2115-2124, 2016. © 2016 Wiley Periodicals, Inc.

HIV-1 early and late diagnosis in the Emilia Romagna Region (Italy): a three year study.

It is crucial to establish the timing of infection and distinguish between early and long-lasting HIV-1 infections not only for partner notification and epidemiological surveillance, but also to offer early drug treatment and contain the spread of infection. This study analyzed serum and/or plasma samples with a first positive HIV antibody/antigen result coming from different Medical Centers in the Emilia Romagna Region, North East Italy, using the avidity assay, Western Blotting, RNA viral load, CD4 cell counts and genotyping assay. From May 2013 to May 2016, we certified 845 new HIV-1 infections, 18.7% of which were classified on the basis of avidity index as recent infections and 81.3% as long-lasting infections, with an estimated conversion time exceeding six months at the time of study. Western Blotting showed reactivity to only one or two HIV-1 proteins in recently infected patients (RIPs), while a complete pattern to gag, env and pol proteins was observed in most long-lasting infected patients (LLIPs). The median age, gender, nationality and risk transmission factors were comparable in RIPs and LLIPs. Phylogenetic analysis performed in available plasma disclosed B strains, non-B subtypes and circulating recombinant forms (CRFs) in both groups of patients, with a major presence of CRFs in non-Italian HIV subjects. The large number of patients unaware of their HIV status makes it crucial to discover hidden epidemics and implement appropriate targeted public health interventions.

Atazanavir and lopinavir profile in pregnant women with HIV: tolerability, activity and pregnancy outcomes in an observational national study.

BACKGROUND: Atazanavir and lopinavir represent the main HIV protease inhibitors recommended in pregnancy, but comparative data in pregnant women are limited. METHODS: Women from a national observational study, exposed in pregnancy to either atazanavir or lopinavir, were compared for glucose and lipid profiles, liver function tests, CD4 count, HIV RNA and main pregnancy outcomes. Statistical methods included univariate and multivariable analyses. RESULTS: The study population included 428 pregnancies (lopinavir, 322; atazanavir, 106). The lopinavir group was characterized by higher rates of HIV diagnosis in pregnancy and treatment indication for maternal health, lower CD4 counts, higher HIV RNA levels, less frequent antiretroviral treatment at conception and shorter duration of drug exposure during pregnancy. No differences in pregnancy outcomes, glucose metabolism and weight gain were observed. The two groups also showed in a multivariable analysis similar odds for detectable HIV RNA in the third trimester (adjusted OR 0.85, 95% CI 0.35-2.10, P = 0.730). Total lipid levels were significantly higher in the lopinavir group (median values in the third trimester 239 versus 221 mg/dL for total cholesterol and 226 versus 181 mg/dL for triglycerides; P < 0.001 for both comparisons) and bilirubin levels were significantly higher in the atazanavir group (1.53 versus 0.46 mg/dL, P < 0.001). CONCLUSIONS: In this observational study atazanavir and lopinavir showed similar safety and activity in pregnancy, with no differences in the main pregnancy outcomes. Atazanavir use was associated with a better lipid profile and with higher bilirubin levels. Overall, the study findings confirm that these two HIV protease inhibitors represent equally valid alternative options.

Relationship between health-related quality of life measures and high HIV viral load in HIV-infected triple-class-experienced patients.

BACKGROUND: Health-related quality of life (HRQoL) has been recognized as a central measure of the overall health status in HIV patients. With the availability of different highly effective drug combinations, maximizing quality-adjusted survival has become a major target of HIV treatment. Although the association of HIV RNA and CD4 cell count with clinical HIV progression has been well established, the relation between these markers and HRQoL measures is still unclear. METHOD: This cross-sectional study investigated the relationship linking HIV RNA and CD4 to HRQoL measures in 181 triple-class-experienced patients with advanced HIV disease. The instrument used was the ISSQoL, a self-administered and HIV-specific HRQoL questionnaire. RESULTS: Data showed no correlation between HRQoL measures and CD4 counts. Higher HIV RNA levels were, however, associated with poor HRQoL scores in 3 out of 9 scales of social functioning, depression and anxiety, and satisfaction with quality of life. In multivariable analyses, only the satisfaction with quality of life mean score remained significantly lower for the HIV RNA ≯100,000 copies/mL group compared to the HIV RNA 50 to 10,000 copies/mL group. CONCLUSIONS: Although other determinants of HRQoL in people with HIV should also be considered, this finding suggests a negative impact of high viral load on perceived HRQoL that adds to other described determinants of lower quality of life in people with HIV, such as lower social support and self-reported symptoms.

Calcium phosphate cement augmentation after volar locking plating of distal radius fracture significantly increases stability.

PURPOSE: Distal radius fractures represent the most common fractures in adults. Volar locking plating to correct unstable fractures has become increasingly popular. Although reasonable primary reduction is possible in most cases, maintenance of reduction until the fracture is healed is often problematic in osteoporotic bone. To our knowledge, no biomechanical studies have compared the effect of enhancement with biomaterial on two different volar fixed-angle plates. METHODS: Human fresh-frozen cadaver pairs of radii were used to simulate an AO/OTA 23-A3 fracture. In a total of four groups (n = 7 for each group), two volar fixed-angle plates (Aptus 2.5 mm locking fracture plate, Medartis, Switzerland and VA-LCP two-column distal radius plate 2.4, volar, Synthes, Switzerland) with or without an additional injection of a biomaterial (Hydroset Injectable HA Bone Substitute, Stryker, Switzerland) into the dorsal comminution zone were used to fix the distal metaphyseal fragment. Each specimen was tested load-controlled under cyclic loading with a servo-hydraulic material testing machine. Displacement, stiffness, dissipated work and failure mode were recorded. RESULTS: Improved mechanical properties (decreased displacement, increased stiffness, decreased dissipated work) were found in both plates if the biomaterial was additionally injected. Improvement of mechanical parameters after biomaterial injection was more evident in the Synthes plate compared to the Aptus plate. Pushing out of the screws was noticed as a failure mode only in samples lacking supplementary biomaterial. CONCLUSIONS: Injection of a biomaterial into the dorsal comminution zone increases stability after volar locking plating of distal radius fractures in vitro.

Response to raltegravir-based salvage therapy in HIV-infected patients with hepatitis C virus or hepatitis B virus coinfection.

OBJECTIVES: To define the impact of coinfection with hepatitis B virus (HBV) or hepatitis C virus (HCV) on viroimmunological response to raltegravir-based salvage regimens that also include new HIV inhibitors such as maraviroc, darunavir and etravirine. METHODS: We used data from a national observational study of patients starting raltegravir-based regimens to compare virological suppression and CD4 cell change from baseline in patients with and without concomitant HBV or HCV infection. RESULTS: Overall, 275 patients (107 coinfected and 168 non-coinfected) were evaluated. Coinfected patients were more commonly former intravenous drug users and had a longer history of HIV infection and higher baseline aminotransferase levels. Both HIV-RNA and CD4 response were similar in the two groups. Mean time to first HIV-RNA copy number <50 copies/mL was 4.1 months (95% CI 3.5-4.6) in non-coinfected patients and 3.9 months (95% CI 3.3-4.5) in coinfected patients (hazard ratio 1.039, 95% CI 0.761-1.418, P = 0.766, log-rank test). The risk of developing new grade 3-4 hepatic adverse events was significantly higher in coinfected patients (hazard ratio 1.779, 95% CI 1.123-2.817, P = 0.009). The two groups of coinfected and non-coinfected patients had similar rates of interruption of any baseline drug (hazard ratio 1.075, 95% CI 0.649-1.781, P = 0.776) and of raltegravir (hazard ratio 1.520, 95% CI 0.671-3.447, P = 0.311). Few AIDS-defining events and deaths occurred. CONCLUSIONS: Viroimmunological response to regimens based on raltegravir and other recent anti-HIV inhibitors is not negatively affected by coinfection with HBV or HCV. Liver toxicity, either pre-existing or new, is more common in coinfected patients, but with no increased risk of treatment interruption.

Pregnancy outcomes in HIV-infected women of advanced maternal age.

BACKGROUND: There is limited information on pregnancy outcomes in women with HIV who are of a more advanced maternal age. METHODS: Data from a national observational study in Italy were used to evaluate the risk of nonelective cesarean section, preterm delivery, low birthweight, major birth defects, and small gestational age-adjusted birthweight according to maternal age (<35 and ≥35 years, respectively). RESULTS: Among 1,375 pregnancies with live births, 82.4% of deliveries were elective cesarean sections, 15.8% were nonelective cesarean sections, and 1.8% were vaginal deliveries. Rates of nonelective cesarean section were similar among mothers ≥35 and <35 years (odds ratio [OR], 1.22; 95% CI, 0.90-1.65;P = .19). Preterm delivery and low birthweight were significantly more common among women ≥35 years in univariate but not in multivariate analyses. Newborns from women ≥35 and <35 years showed no differences inZ scores of birthweight, with a similar occurrence of birthweight <10th percentile (12.1% vs 12.0%; OR, 1.02; 95% CI, 0.71-1.46;P = .93). The overall rate of birth defects was 3.4% (95% CI, 2.4-4.4), with no differences by maternal age (≥35 years, 3.5%; <35 years, 3.3%; OR, 1.05; 95% CI, 0.56-1.98;P = .88). DISCUSSION: In this study of pregnant women with HIV, older women were at higher risk of some adverse pregnancy outcomes, such as preterm delivery and low birthweight. The association, however, did not persist in multivariable analyses, suggesting a role of some predisposing factors associated with older age.

Current concepts review: Management of civilian transpelvic gunshot fractures.

INTRODUCTION: Civilian gunshot fractures of the pelvic ring represent a unique challenge for orthopaedic surgeons due to a high incidence of complicating associated injuries. Internationally accepted guidelines for these injuries are not available. The aim of this review is to summarize the available literature and to provide concise management recommendations. METHODS: Literature search was performed using PubMed. The review focuses on civilian gunshot fractures of the pelvic ring and includes the acetabulum and hip joint only where it was deemed necessary for the understanding of the management of these patients. RESULTS: The management of civilian transpelvic gunshot fractures is complicated by potentially life-threatening associated injuries, the risk of contamination with bowel content and retained bullets in joints. The infection risk is higher compared to extremity gunshot fractures. There is no clear evidence for the use of antibiotics available. The studies focusing on civilian pelvic ring gunshot fractures reported no case of orthopaedic fracture fixation in their series. Routine wash-out and debridement of fractures is not warranted based on the literature but conflicting recommendations for surgical interventions exist. CONCLUSION: There is limited evidence available for civilian transpelvic gunshot fractures. The high frequency of associated injuries requires a thorough clinical examination and multidisciplinary management. We recommend routine antibiotic prophylaxis for all transpelvic gunshots. For fractures with a high risk of infection, a minimum of 24 h broad-spectrum antibiotics is recommended. The indication for orthopaedic fixation of civilian transpelvic gunshot fractures is based on the assessment of the stability of the fracture and is rarely necessary. Although conflicting recommendations exist, routine wash-out and debridement is not recommended based on the literature. Bullets buried in bone without contact to synovial fluid do not warrant removal, unless they have traversed large bowel and are accessible without undue morbidity. Furthermore, bullets should be routinely removed if they are retained in the hip joint, if mechanical irritation of soft tissues by projectiles is expected or if the bullet traversed large bowel before entering the hip joint.

The Hidden Pandemic: Impact of the COVID-19 Pandemic on Trauma Cases Due to Domestic Violence Admitted to the Biggest Level-One Trauma Center in Austria.

BACKGROUND: An alarming increase in domestic violence was reported during the COVID-19 pandemic worldwide. The aim of this study is to investigate changes in the frequency and the nature of domestic violence at the largest level-one trauma center in Austria. METHODS: All patients admitted to our institution with domestic violence injuries 15 months before and after the beginning of the COVID-19 pandemic were included. For our analysis, we investigated the frequency of trauma patients after domestic violence in relation to all other trauma patients. Furthermore, age, sex, citizenship, injury pattern, injured body regions, injury mechanism, offender-victim relationship, and hospitalization rate were also analyzed. RESULTS: Among all trauma patients admitted, the ratio of patients who reported domestic violence injuries increased from 0.465% to 0.548% since the start of the pandemic. In addition, out of the total count of domestic violence victims, the percentage of Austrian citizens increased significantly from 51.2% to 60.6% (p = 0.016). All other parameters showed no significant changes pre and post-pandemic. CONCLUSION: The COVID-19 pandemic contributed to a relative increase in patients with domestic violence injuries at the largest trauma unit in Austria, along with a significant increase among Austrian citizens. The remaining study parameters did not differ significantly, indicating that the frequency changed during the pandemic but not the underlying pattern of domestic violence.

The importance of additional intracranial injuries in epidural hematomas: detailed clinical analysis, long-term outcome, and literature review in surgically managed epidural hematomas.

Objective: Epidural hematomas (EDH) occur in up to 8.2% of all traumatic brain injury patients, with more than half needing surgical treatment. In most patients suffering from this perilous disease, good recovery with an excellent clinical course is possible. However, the clinical course is mainly dependent on the presence of additional intracerebral injuries. Few studies comparing isolated and combined EDH in detail exist. Methods: We performed a retrospective single-center study from April 2002 to December 2014. The mean follow-up time was more than 6 years. In addition to analyzing diverse clinicoradiological data, we performed a systematic literature review dealing with a detailed comparison of patients with (combined) and without (isolated) additional intracerebral injuries. Results: We included 72 patients in the study. With increasing age, combined EDH had a higher incidence than isolated EDH. The mortality rate of the patients in the cohort was 10%, of which 0% had isolated EDH and 10% had combined EDH. Good recovery was achieved in 69% of patients, of which 91% had isolated EDH and 50% had combined EDH. A subgroup analysis of the different additional intracerebral injuries in combined EDH demonstrated no significant difference in outcome. A systematic literature review only identified six studies. Patients with isolated EDH had a statistically significantly lower mortality risk [relative risk (RR): 0.22; 95% CI: 0.12-0.39] and a statistically significantly lower risk of unfavorable Glasgow outcome scale score (RR: 0.21; 95% CI: 0.14-0.31) than patients with combined EDH. Conclusions: An excellent outcome in patients with surgically treated isolated EDH is possible. Furthermore, patients with combined EDH or isolated EDH with a low Glasgow coma scale (GCS) score may have favorable outcomes in 50% of the cases. Therefore, every possible effort for treatment should be made for this potentially lethal injury.

Evolution of proviral DNA HIV-1 tropism under selective pressure of maraviroc-based therapy.

OBJECTIVES: To evaluate the evolution of HIV-1 coreceptor tropism in proviral DNA of patients during maraviroc-based therapy. METHODS: Fourteen heavily high active antiretroviral therapy (HAART)-treated patients with a CCR5 Trofile profile were monitored over a 24 month period from the start of maraviroc therapy. Whole-blood samples were obtained at different timepoints, and coreceptor tropism was determined for proviral DNA from the V3-loop region sequence using the Geno2Pheno algorithm [false positive rate (FPR): 20%]. RESULTS: At the start of maraviroc treatment, 13/14 patients were viraemic (median: 4.33 log copies/mL). Concordance in R5 tropism (R5/R5) was observed between circulating HIV-RNA (Trofile) and HIV-DNA provirus in 10/14 patients (median FPR = 54.0%), while 4 patients showed a CXCR4-tropic R5/X4 variant in their provirus (FPR: 5.8%, 5.7%, 16.6% and 1.1%, respectively). All R5/R5 patients showed a stable HIV-1 DNA coreceptor usage. Two out of four R5/X4 patients showed a tropism shift in their archived provirus and, after 6 months a prevalence of R5-tropic virus was detected in DNA. The other two R5/X4 patients harboured the 11/25 genotype, and maintained X4 tropism in provirus during the study. Virological response did not reveal differences in RNA decay and CD4+ cell recovery in patients with discordant tropism. CONCLUSIONS: A relatively good correlation between RNA and DNA tropism was observed at baseline. Proviral DNA tropism remained stable over 24 months of maraviroc-based therapy, indicating that determination of proviral DNA V3 sequence could be used in tropism prediction in clinical practice. The data also confirm the importance of the 11/25 rule in predicting viral tropism.

Assessment of a novel biomechanical fracture model for distal radius fractures.

BACKGROUND: Distal radius fractures (DRF) are one of the most common fractures and often need surgical treatment, which has been validated through biomechanical tests. Currently a number of different fracture models are used, none of which resemble the in vivo fracture location. The aim of the study was to develop a new standardized fracture model for DRF (AO-23.A3) and compare its biomechanical behavior to the current gold standard. METHODS: Variable angle locking volar plates (ADAPTIVE, Medartis) were mounted on 10 pairs of fresh-frozen radii. The osteotomy location was alternated within each pair (New: 10 mm wedge 8 mm / 12 mm proximal to the dorsal / volar apex of the articular surface; Gold standard: 10 mm wedge 20 mm proximal to the articular surface). Each specimen was tested in cyclic axial compression (increasing load by 100 N per cycle) until failure or -3 mm displacement. Parameters assessed were stiffness, displacement and dissipated work calculated for each cycle and ultimate load. Significance was tested using a linear mixed model and Wald test as well as t-tests. RESULTS: 7 female and 3 male pairs of radii aged 74 ± 9 years were tested. In most cases (7/10), the two groups showed similar mechanical behavior at low loads with increasing differences at increasing loads. Overall the novel fracture model showed a significant different biomechanical behavior than the gold standard model (p < 0,001). The average final loads resisted were significantly lower in the novel model (860 N ± 232 N vs. 1250 N ± 341 N; p = 0.001). CONCLUSION: The novel biomechanical fracture model for DRF more closely mimics the in vivo fracture site and shows a significantly different biomechanical behavior with increasing loads when compared to the current gold standard.