The spatio-temporal dynamics of PKA activity profile during mitosis and its correlation to chromosome segregation

The cyclic adenosine monophosphate dependent kinase protein (PKA) controls a variety of cellular processes including cell cycle regulation. Here, we took advantages of genetically encoded FRET-based biosensors, using an AKAR-derived biosensor to characterize PKA activity during mitosis in living HeLa cells using a single-cell approach. We measured PKA activity changes during mitosis. HeLa cells exhibit a substantial increase during mitosis, which ends with telophase. An AKAREV T>A inactive form of the biosensor and H89 inhibitor were used to ascertain for the specificity of the PKA activity measured. On a spatial point of view, high levels of activity near to chromosomal plate during metaphase and anaphase were detected. By using the PKA inhibitor H89, we assessed the role of PKA in the maintenance of a proper division phenotype. While this treatment in our hands did not impaired cell cycle progression in a drastic manner, inhibition of PKA leads to a dramatic increase in chromososme misalignement on the spindle during metaphase that could result in aneuploidies. Our study emphasizes the insights that can be gained with genetically encoded FRET-based biosensors, which enable to overcome the shortcomings of classical methologies and unveil in vivo PKA spatiotemporal profiles in HeLa cells.     

Computed tomography-based joint locations affect calculation of joint moments during gait when compared to scaling approaches

Hip joint moments are an important parameter in the biomechanical evaluation of orthopaedic surgery. Joint moments are generally calculated using scaled generic musculoskeletal models. However, due to anatomical variability or pathology, such models may differ from the patient's anatomy, calling into question the accuracy of the resulting joint moments. This study aimed to quantify the potential joint moment errors caused by geometrical inaccuracies in scaled models, during gait, for eight test subjects. For comparison, a semi-automatic computed tomography (CT)-based workflow was introduced to create models with subject-specific joint locations and inertial parameters. 3D surface models of the femora and hemipelves were created by segmentation and the hip joint centres and knee axes were located in these models. The scaled models systematically located the hip joint centre (HJC) up to 33.6 mm too inferiorly. As a consequence, significant and substantial peak hip extension and abduction moment differences were recorded, with, respectively, up to 23.1% and 15.8% higher values in the image-based models. These findings reaffirm the importance of accurate HJC estimation, which may be achieved using CT- or radiography-based subject-specific modelling. However, obesity-related gait analysis marker placement errors may have influenced these results and more research is needed to overcome these artefacts.     

Subject-specific hip geometry and hip joint centre location affects calculated contact forces at the hip during gait

Hip loading affects the development of hip osteoarthritis, bone remodelling and osseointegration of implants. In this study, we analyzed the effect of subject-specific modelling of hip geometry and hip joint centre (HJC) location on the quantification of hip joint moments, muscle moments and hip contact forces during gait, using musculoskeletal modelling, inverse dynamic analysis and static optimization. For 10 subjects, hip joint moments, muscle moments and hip loading in terms of magnitude and orientation were quantified using three different model types, each including a different amount of subject-specific detail: (1) a generic scaled musculoskeletal model, (2) a generic scaled musculoskeletal model with subject-specific hip geometry (femoral anteversion, neck-length and neck-shaft angle) and (3) a generic scaled musculoskeletal model with subject-specific hip geometry including HJC location. Subject-specific geometry and HJC location were derived from CT. Significant differences were found between the three model types in HJC location, hip flexion–extension moment and inclination angle of the total contact force in the frontal plane. No model agreement was found between the three model types for the calculation of contact forces in terms of magnitude and orientations, and muscle moments. Therefore, we suggest that personalized models with individualized hip joint geometry and HJC location should be used for the quantification of hip loading. For biomechanical analyses aiming to understand modified hip joint loading, and planning hip surgery in patients with osteoarthritis, the amount of subject-specific detail, related to bone geometry and joint centre location in the musculoskeletal models used, needs to be considered.     

Semi-automated segmentation and visualisation of outer bone cortex from medical images

Good segmentation of the outer bone cortex from medical images is a prerequisite for applications in the field of finite element analysis, surgical planning environments and personalised, case dependent, bone reconstruction. However, current segmentation procedures are often unsatisfactory. This study presents an automated filter procedure to generate a set of adapted contours from which a surface mesh can be deduced directly. The degree of interaction is user determined. The bone contours are extracted from the patients CT data by quick grey value segmentation. An extended filter procedure then only retains contour information representing the outer cortex as more specific internal loops and shape irregularities are removed, tailoring the image for the above-mentioned applications. The developed medical image based design methodology to convert contour sets of multiple bone types, from tibia tumour to neurocranium, is reported and discussed.     

Dual Effects of Hydrogen Sulfide Donor on Meiosis and Cumulus Expansion of Porcine Cumulus-Oocyte Complexes

Hydrogen sulfide (H₂S) has been revealed to be a signal molecule with second messenger action in the somatic cells of many tissues, including the reproductive tract. The aim of this study was to address how exogenous H₂S acts on the meiotic maturation of porcine oocytes, including key maturation factors such as MPF and MAPK, and cumulus expansion intensity of cumulus-oocyte complexes. We observed that the H₂S donor, Na₂S, accelerated oocyte in vitro maturation in a dose-dependent manner, following an increase of MPF activity around germinal vesicle breakdown. Concurrently, the H₂S donor affected cumulus expansion, monitored by hyaluronic acid production. Our results suggest that the H₂S donor influences oocyte maturation and thus also participates in the regulation of cumulus expansion. The exogenous H₂S donor apparently affects key signal pathways of oocyte maturation and cumulus expansion, resulting in faster oocyte maturation with little need of cumulus expansion.