Development and performance evaluation of a multi-PID muscle loading driven in vitro active-motion shoulder simulator and application to assessing reverse total shoulder arthroplasty.

In vitro active shoulder motion simulation can provide improved understanding of shoulder biomechanics; however, accurate simulators using advanced control theory have not been developed. Therefore, our objective was to develop and evaluate a simulator which uses real-time kinematic feedback and closed loop proportional integral differential (PID) control to produce motion. The simulator's ability to investigate a clinically relevant variable-namely muscle loading changes resulting from reverse total shoulder arthroplasty (RTSA)-was evaluated and compared to previous findings to further demonstrate its efficacy. Motion control of cadaveric shoulders was achieved by applying continuously variable forces to seven muscle groups. Muscle forces controlling each of the three glenohumeral rotational degrees of freedom (DOF) were modulated using three independent PID controllers running in parallel, each using measured Euler angles as their process variable. Each PID controller was configured and tuned to control the loading of a set of muscles which, from previous in vivo investigations, were found to be primarily responsible for movement in the PID's DOF. The simulator's ability to follow setpoint profiles for abduction, axial rotation, and horizontal extension was assessed using root mean squared error (RMSE) and average standard deviation (ASD) for multiple levels of arm mass replacement. A specimen was then implanted with an RTSA, and the effect of joint lateralization (0, 5, 10 mm) on the total deltoid force required to produce motion was assessed. Maximum profiling error was <2.1 deg for abduction and 2.2 deg for horizontal extension with RMSE of <1 deg. The nonprofiled DOF were maintained to within 5.0 deg with RMSE <1.0 deg. Repeatability was high, with ASDs of <0.31 deg. RMSE and ASD were similar for all levels of arm mass replacement (0.73-1.04 and 0.14-0.22 deg). Lateralizing the joint's center of rotation (CoR) increased total deltoid force by up to 8.5% body weight with the maximum early in abduction. This simulator, which is the first to use closed loop control, accurately controls the shoulder's three rotational DOF with high repeatability, and produces results that are in agreement with previous investigations. This simulator's improved performance, in comparison to others, increases the statistical power of its findings and thus its ability to provide new biomechanical insights.

The tobacco MAP215/Dis1-family protein TMBP200 is required for the functional organization of microtubule arrays during male germline establishment.

The haploid microspore division during pollen development in flowering plants is an intrinsically asymmetric division which establishes the male germline for sexual reproduction. Arabidopsis gem1 mutants lack the male germline as a result of disturbed microspore polarity, division asymmetry, and cytokinesis and represent loss-of-function mutants in MOR1/GEM1, a plant orthologue of the conserved MAP215/Dis1 microtubule associated protein (MAP) family. This provides genetic evidence for the role of MAP215/Dis1 in the organization of gametophytic microtubule arrays, but it has remained unknown how microtubule arrays are affected in gem1 mutant microspores. Here, novel male gametophytic microtubule-reporter Nicotiana tabacum plants were constructed, expressing a green fluorescent protein-alpha-TUBULIN fusion protein (GFP-TUA6) under the control of a microspore-specific promoter. These plants allow effective visualization of all major male gametophytic microtubule arrays and provide useful tools to study the regulation of microtubule arrays by MAPs and other effectors. Depletion of TMBP200, a tobacco homologue of MOR1/GEM1 in gametophytic microtubule-reporter plants using microspore-targeted RNA interference, induced defects in microspore polarity, division asymmetry and cytokinesis that were associated with striking defects in phragmoplast position, orientation, and structure. Our observations further reveal a requirement for TMBP200 in gametophytic spindle organization and a novel role in spindle position and orientation in polarized microspores. These results provide direct evidence for the function of MAP215/Dis1 family protein TMBP200 in the organization of microtubule arrays critical for male germline formation in plants.

Identification of microspore-active promoters that allow targeted manipulation of gene expression at early stages of microgametogenesis in Arabidopsis.

BACKGROUND: The effective functional analysis of male gametophyte development requires new tools enabling the spatially and temporally controlled expression of both marker genes and modified genes of interest. In particular, promoters driving expression at earlier developmental stages including microspores are required. RESULTS: Transcriptomic datasets covering four progressive stages of male gametophyte development in Arabidopsis were used to select candidate genes showing early expression profiles that were male gametophyte-specific. Promoter-GUS reporter analysis of candidate genes identified three promoters (MSP1, MSP2, and MSP3) that are active in microspores and are otherwise specific to the male gametophyte and tapetum. The MSP1 and MSP2 promoters were used to successfully complement and restore the male transmission of the gametophytic two-in-one (tio) mutant that is cytokinesis-defective at first microspore division. CONCLUSION: We demonstrate the effective application of MSP promoters as tools that can be used to elucidate gametophytic gene functions in microspores in a male-specific manner.