Kinetochore-spindle microtubule interactions during mitosis.

The kinetochore is a proteinaceous structure that assembles onto centromeric DNA and mediates chromosome attachment to microtubules during mitosis. This description is deceivingly simple: recent proteomic studies suggest that the diminutive kinetochores of Saccharomyces cerevisiae are comprised of at least 60 proteins organized into as many as 14 different subcomplexes. Many of these proteins, such as the centromeric histone variant CENP-A, and entire subcomplexes, such as the Ndc80(Hec1) complex, are conserved from yeast to humans despite the diverse nature of the DNA sequences on which they assemble. There have recently been advances in our understanding of the molecular basis of how kinetochores establish dynamic attachments to spindle microtubules, and how these attachments are correctly oriented to ensure segregation of sister chromatids to daughter cells.

Estimating insertion length of umbilical arterial and venous catheters in newborn infants: time for change.

BACKGROUND: Umbilical catheters are inserted through the umbilical artery or vein at birth and are crucial in neonatal care. There are several different methods of estimating adequate insertion length of umbilical catheters based on one of two hypotheses; that the insertion length of the UC is correlated to either the infant's birth weight or an external length measurement. AIM: To review the published literature on methods of estimating insertion lengths of umbilical arterial catheters (UACs) and umbilical venous catheters (UVCs) in newborn infants. METHODS: Systematic search on Medline was undertaken using keywords for relevant papers up to March 2019. Papers were selected by manual search of titles and abstracts. RESULTS: Formulae for predicting umbilical catheter insertion length are unreliable, particularly for UVCs. There is also conflicting evidence around whether birth weight-based formulae are more reliable than external length-based formulae. Studies comparing various methods to determine their efficacy to show that current formulae have a low accuracy for determining both UVC and UAC positioning. CONCLUSIONS: Current formulae for estimating insertion length of umbilical catheters are not fit for purpose. We propose a new observational study which uses a new external length measurement, the sternal notch to umbilicus length, to develop a more reliable formula for the insertion of UVC and UAC to an adequate length.

An Assessment of Research Priorities to Dampen the Pendulum Swing of Burn Resuscitation.

On June 17 to 18, 2019, the American Burn Association, in conjunction with Underwriters Laboratories, convened a group of experts on burn resuscitation in Washington, DC. The goal of the meeting was to identify and discuss novel research and strategies to optimize the process of burn resuscitation. Patients who sustain a large thermal injury (involving >20% of the total body surface area [TBSA]) face a sequence of challenges, beginning with burn shock. Over the last century, research has helped elucidate much of the underlying pathophysiology of burn shock, which places multiple organ systems at risk of damage or dysfunction. These studies advanced the understanding of the need for fluids for resuscitation. The resultant practice of judicious and timely infusion of crystalloids has improved mortality after major thermal injury. However, much remains unclear about how to further improve and customize resuscitation practice to limit the morbidities associated with edema and volume overload. Herein, we review the history and pathophysiology of shock following thermal injury, and propose some of the priorities for resuscitation research. Recommendations include: studying the utility of alternative endpoints to resuscitation, reexamining plasma as a primary or adjunctive resuscitation fluid, and applying information about inflammation and endotheliopathy to target the underlying causes of burn shock. Undoubtedly, these future research efforts will require a concerted effort from the burn and research communities.

Full-length dimeric MCAK is a more efficient microtubule depolymerase than minimal domain monomeric MCAK.

MCAK belongs to the Kinesin-13 family, whose members depolymerize microtubules rather than translocate along them. We defined the minimal functional unit of MCAK as the catalytic domain plus the class specific neck (MD-MCAK), which is consistent with previous reports. We used steady-state ATPase kinetics, microtubule depolymerization assays, and microtubule.MCAK cosedimentation assays to compare the activity of full-length MCAK, which is a dimer, with MD-MCAK, which is a monomer. Full-length MCAK exhibits higher ATPase activity, more efficient microtubule end binding, and reduced affinity for the tubulin heterodimer. Our studies suggest that MCAK dimerization is important for its catalytic cycle by promoting MCAK binding to microtubule ends, enhancing the ability of MCAK to recycle for multiple rounds of microtubule depolymerization, and preventing MCAK from being sequestered by tubulin heterodimers.

Mustard gas and American race-based human experimentation in World War II.

This essay examines the risks of racialized science as revealed in the American mustard gas experiments of World War II. In a climate of contested beliefs over the existence and meanings of racial differences, medical researchers examined the bodies of Japanese American, African American, and Puerto Rican soldiers for evidence of how they differed from whites.

The microtubule-destabilizing kinesin XKCM1 is required for chromosome positioning during spindle assembly.

Xenopus kinesin catastrophe modulator-1 (XKCM1) is a Kin I kinesin family member that uses the energy of ATP hydrolysis to depolymerize microtubules. We demonstrated previously that XKCM1 is essential for mitotic-spindle assembly in vitro and acts by regulating microtubule dynamics as a pure protein, in extracts and in cells. A portion of the XKCM1 pool is specifically localized to centromeres during mitosis and may be important in chromosome movement. To selectively analyze the function of centromere-bound XKCM1, we generated glutathione-S-transferase (GST) fusion proteins containing the N-terminal globular domain (GST-NT), the centrally located catalytic domain (GST-CD), and the C-terminal alpha-helical tail (GST-CT) of XKCM1. The GST-NT protein targeted to centromeres during spindle assembly, suggesting that the N-terminal domain of XKCM1 is sufficient for centromere localization. Addition of GST-NT prior to or after spindle assembly replaced endogenous XKCM1, indicating that centromere targeting is a dynamic process. Loss of endogenous XKCM1 from centromeres caused a misalignment of chromosomes on the metaphase plate without affecting global spindle structure. These results suggest that centromere bound XKCM1 has an important role in chromosome positioning on the spindle.

Aurora B phosphorylates centromeric MCAK and regulates its localization and microtubule depolymerization activity.

BACKGROUND: Sister kinetochores must bind microtubules in a bipolar fashion to equally segregate chromosomes during mitosis. The molecular mechanisms underlying this process remain unclear. Aurora B likely promotes chromosome biorientation by regulating kinetochore-microtubule attachments. MCAK (mitotic centromere-associated kinesin) is a Kin I kinesin that can depolymerize microtubules. These two proteins both localize to mitotic centromeres and have overlapping mitotic functions, including regulation of microtubule dynamics, proper chromosome congression, and correction of improper kinetochore-microtubule attachments. RESULTS: We show that Aurora B phosphorylates and regulates MCAK both in vitro and in vivo. Specifically, we mapped six Aurora B phosphorylation sites on MCAK in both the centromere-targeting domain and the neck region. Aurora B activity was required to localize MCAK to centromeres, but not to spindle poles. Aurora B phosphorylation of serine 196 in the neck region of MCAK inhibited its microtubule depolymerization activity. We found that this key site was phosphorylated at centromeres and anaphase spindle midzones in vivo. However, within the inner centromere there were pockets of both phosphorylated and unphosphorylated MCAK protein, suggesting that phosphate turnover is crucial in the regulation of MCAK activity. Addition of alpha-p-S196 antibodies to Xenopus egg extracts or injection of alpha-p-S196 antibodies into cells caused defects in chromosome positioning and/or segregation. CONCLUSIONS: We have established a direct link between the microtubule depolymerase MCAK and Aurora B kinase. Our data suggest that Aurora B both positively and negatively regulates MCAK during mitosis. We propose that Aurora B biorients chromosomes by directing MCAK to depolymerize incorrectly oriented kinetochore microtubules.

A comparison of writing exercises to motivate young women to practise breast self-examinations.

OBJECTIVES: This study investigated the novel use of written emotional disclosure procedures for motivating health behavior change, specifically engagement in breast self examination (BSE) among college-aged women. Writing instructions were tailored to a precontemplative stage of change in accordance with the transtheoretical model proposed by Prochaska and DiClemente (1983) to test the hypothesis that stage-matched writing strategy would facilitate greater contemplation than a stage-mismatched writing strategy and a control condition. DESIGN AND METHODS: Precontemplative women (N=55) were randomly assigned to one of three groups: (a) a stage-matched consciousness-raising writing exercise; (b) a stage-mismatched BSE action-oriented exercise; or (c) a control condition, and measured on contemplation, preparation, and action, as well as performance of BSEs. RESULTS: Analyses indicated that both the consciousness-raising writing exercise and the BSE action-oriented exercise increased participants' level of contemplation when compared to the control condition. Women assigned to the BSE action-oriented exercise endorsed more statements indicative of an action stage of change and exhibited a trend for greater performance of BSE compared to the other two conditions. CONCLUSIONS: The application of writing exercises and the relevance of stage-matching is discussed in the context of the stage of change model.

The microtubule-destabilizing kinesin XKCM1 regulates microtubule dynamic instability in cells.

The dynamic activities of cellular microtubules (MTs) are tightly regulated by a balance between MT-stabilizing and -destabilizing proteins. Studies in Xenopus egg extracts have shown that the major MT destabilizer during interphase and mitosis is the kinesin-related protein XKCM1, which depolymerizes MT ends in an ATP-dependent manner. Herein, we examine the effects of both overexpression and inhibition of XKCM1 on the regulation of MT dynamics in vertebrate somatic cells. We found that XKCM1 is a MT-destabilizing enzyme in PtK2 cells and that XKCM1 modulates cellular MT dynamics. Our results indicate that perturbation of XKCM1 levels alters the catastrophe frequency and the rescue frequency of cellular MTs. In addition, we found that overexpression of XKCM1 or inhibition of KCM1 during mitosis leads to the formation of aberrant spindles and a mitotic delay. The predominant spindle defects from excess XKCM1 included monoastral and monopolar spindles, as well as small prometaphase-like spindles with improper chromosomal attachments. Inhibition of KCM1 during mitosis led to prometaphase spindles with excessively long MTs and spindles with partially separated poles and a radial MT array. These results show that KCM1 plays a critical role in regulating both interphase and mitotic MT dynamics in mammalian cells.

Depletion of centromeric MCAK leads to chromosome congression and segregation defects due to improper kinetochore attachments.

The complex behavior of chromosomes during mitosis is accomplished by precise binding and highly regulated polymerization dynamics of kinetochore microtubules. Previous studies have implicated Kin Is, unique kinesins that depolymerize microtubules, in regulating chromosome positioning. We have characterized the immunofluorescence localization of centromere-bound MCAK and found that MCAK localized to inner kinetochores during prophase but was predominantly centromeric by metaphase. Interestingly, MCAK accumulated at leading kinetochores during congression but not during segregation. We tested the consequences of MCAK disruption by injecting a centromere dominant-negative protein into prophase cells. Depletion of centromeric MCAK led to reduced centromere stretch, delayed chromosome congression, alignment defects, and severe missegregation of chromosomes. Rates of chromosome movement were unchanged, suggesting that the primary role of MCAK is not to move chromosomes. Furthermore, we found that disruption of MCAK leads to multiple kinetochore-microtubule attachment defects, including merotelic, syntelic, and combined merotelic-syntelic attachments. These findings reveal an essential role for Kin Is in prevention and/or correction of improper kinetochore-microtubule attachments.

Dinner at Orazio's--David Triggle the model of a mentor.

What does it mean to be a mentor in science? Definitions of mentorship are freely spouted in publications and include concepts such as academic support, professional development, role modeling, interaction, impartment of knowledge, evaluation of work, demonstration of methodology, etc. Perhaps most of us would agree with the duties listed. But just what does it mean, for example, to offer academic support? How might one facilitate professional development for a mentee? While we may agree to the general obligations of a mentor the specifics of what these entail would prove more controversial. This article will illustrate how easy it is to pick out a bad mentor. There are certain elements of conduct that, if practiced, undoubtedly put you in the "bad mentor" category. However, it is very difficult to explain just what it means to be not only an adequate mentor but also a stellar one. It may be easy to list the roles and responsibilities of a mentor but just how should they be performed/carried out? David Triggle is the model of an extraordinary mentor. The conclusion of this paper will focus on some specific mentorship activities David Triggle carried out that illustrate some of the intangible aspects of excellence in mentorship.

Twentieth-century social and health-care influences on location of death in Canada.

Providing the right care, in the right place, to dying persons is hampered by a lack of understanding of where death and dying normally take place and ignorance about what influences location of death. This paper reports the findings of a multidisciplinary historical investigation of 20th-century influences on location of death in Canada. It builds on a study that found a hospitalization-of-death trend in Canada over much of the 20th century but a reduction in hospital deaths beginning in 1994. This study found 2 key influences on location of death: (1) health-care and health-system developments that consolidated care in hospitals while also raising and sustaining public expectations of beneficial if not curative hospital care--the rising hospital-death rate throughout the 20th century can thus be considered an outcome of the shift of illness care from the home to the hospital; and (2) reduced availability of home-based caregivers. A number of developments limited the availability of home care for chronically ill and terminally ill persons, including the increased participation of women in the workforce and the shift in nursing from private home duty to hospitals. Although some health and social support for home care has developed recently, this support clearly does not match that for hospital care. These findings indicate that location of death is an important focal point for studying and planning improvements in end-of-life care.

Functional morbidity following latissimus dorsi flap breast reconstruction.

According to the National Institutes of Health, more than 230,000 new cases of breast cancer will be diagnosed in 2014 alone. Following mastectomy, several reconstructive options exist for women with breast cancer. The timing and approach for reconstruction must be addressed rapidly. Although abdominal tissue transfer is described as the preferred method, it may not be best suited to all patients. The latissimus dorsi (LD) muscle flap is a widely available, proven, and reliable modality. The majority of studies support that shoulder functional morbidity is minimal, but this should be more accurately quantified to allow patients to assess the possible impact on their daily lives. A critical appraisal of the available evidence was undertaken to determine the incidence of new functional morbidity involving the ipsilateral arm following LD pedicled flap breast reconstruction. The process for identifying articles included preappraised and secondary literature sources published between 2005 and 2013. Randomized controlled trials, evidence-based practice, clinical guidelines, and systematic reviews were the quality filters applied. This literature review confirmed that LD muscle transfer does lead to measurable reductions in shoulder joint stability, strength, range of motion, and general functionality. However, these deficiencies resolve in the vast majority of women within 6 to 12 months. Ultimately, the consequences of shoulder function morbidity must be considered and discussed with patients prior to making a final decision.

Managing the child with severe primary insulin-like growth factor-1 deficiency (IGFD): IGFD diagnosis and management.

Growth failure associated with severe primary insulin-like growth factor 1 (IGF-1) deficiency (SPIGFD), a condition defined as basal IGF-1 standard deviation score (SDS) less than or equal to -3 and height SDS less than or equal to -3 in a child with normal or elevated levels of growth hormone, can be successfully treated with the recombinant human IGF-1 mecasermin. In this review, we describe the most safe and effective way to use mecasermin in the treatment of patients with SPIGFD, including how to initiate dosing, key side effects, and how to monitor treatment. Finally, mention of how to reinitiate therapy is made, given the recent drug shortage with mecasermin.

Dynamic dose-feedback prostate brachytherapy in patients with large prostates and/or planned transurethral surgery before implantation.

OBJECTIVE: To compare the quality of permanent prostate brachytherapy (PPB) implants, dosimetric outcomes and urinary morbidity between patients with large (>50 mL) and those with smaller prostates, treated with a dynamic dose-feedback technique as monotherapy for localized prostate cancer. PATIENTS AND METHODS: The series included patients with pre-existing bladder outlet obstruction managed with planned transurethral resection or incision of the prostate; 155 consecutive men had PPB implants as monotherapy for localized prostate cancer using a dynamic dose-feedback approach. Dosimetric variables assessed included the implant volume, the minimum dose to 90% of the prostate (D90), and the volumes of prostate receiving 100% and 150% of the prescribed dose as a percentage of the total volume (V100 and V150), during and after implantation. Urinary morbidity was recorded in terms of acute urinary retention (AUR), the need for surgical intervention after implantation and the American Urologic Association (AUA) symptom score at baseline, 1.5, 3, 6, 9, 12 and 18 months. RESULTS: In all, 38 patients had prostate volumes of >or=50 mL; prostate volume had no influence on any dosimetric variable assessed. Two patients with large prostates (>or=50 mL) had AUR and required delayed surgery. Three patients with small prostates (<50 mL) had transient retention; the differences were not statistically significant (Fisher's exact test). AUA symptom scores peaked at 6 weeks and returned to baseline within a year; there were no statistically significant differences between the groups. Eight patients had planned transurethral surgery at >or=4 months before implantation; they all had D90s of >130 Gy and had no incontinence. CONCLUSION: Using the dynamic feedback technique, there was no adverse dosimetric and urinary morbidity in men having PPB and with prostates of >50 mL. Likewise, there were no impediments, e.g. pubic arch interference, which precluded a favourable dosimetric implant in men with a large prostate. Large prostates should not be a contraindication to PPB and require no hormonal cytoreduction. Patients with obstructive lower urinary tract symptoms can be managed with planned transurethral prostatic surgery before implantation, without compromising implant quality or morbidity.