The detyrosination/re-tyrosination cycle of tubulin and its role and dysfunction in neurons and cardiomyocytes.

Among the variety of post-translational modifications to which microtubules are subjected, the detyrosination/re-tyrosination cycle is specific to tubulin. It is conserved by evolution and characterized by the enzymatic removal and re-addition of a gene-encoded tyrosine residue at the C-terminus of α-tubulin. Detyrosinated tubulin can be further converted to Δ2-tubulin by the removal of an additional C-terminal glutamate residue. Detyrosinated and Δ2-tubulin are carried by stable microtubules whereas tyrosinated microtubules are present on dynamic polymers. The cycle regulates trafficking of many cargo transporting molecular motors and is linked to the microtubule dynamics via regulation of microtubule interactions with specific cellular effectors such as kinesin-13. Here, we give an historical overview of the general features discovered for the cycle. We highlight the recent progress toward structure and functioning of the enzymes that keep the levels of tyrosinated and detyrosinated tubulin in cells, the long-known tubulin tyrosine ligase and the recently discovered vasohibin-SVBP complexes. We further describe how the cycle controls microtubule functions in healthy neurons and cardiomyocytes and how deregulations of the cycle are involved in dysfunctions of these highly differentiated cells, leading to neurodegeneration and heart failure in humans.

Impact of sex and age on the lateralisation of the tibial tubercle in normal paediatric and adolescent populations.

PURPOSE: Numerous methods have been proposed to characterise tubercle lateralisation. However, their normal values and related changes remain unclear. Accordingly, it was aimed to determine the potential sex and age effects and determined the optimal individualised method of diagnosing lateralisation of the tibial tubercle in patients with recurrent patellar dislocation (RPD). METHODS: Measurements included the tibial tubercle-trochlear groove (TT-TG) distance, tibial tubercle-posterior cruciate ligament (TT-PCL) distance and tibial tubercle lateralisation (TTL); and the proximal tibial width (PTW), trochlear width (TW) and trochlear dysplasia index (TDI), for adjustment. A two-way analysis of variance was used to determine the effect of age, sex and their interaction within the normal group. When the age effect was statistically significant, a nonlinear regression was created. Areas under the receiver-operating characteristic curve (AUCs) were calculated to assess diagnostic accuracy. RESULTS: A total of 277 normal participants (mean [SD] age, 13.5 [2.6] years; 125 [45.1%] female) and 227 patients with RPD (mean [SD] age, 13.5 [2.6] years; 161 [58.1%] female) were analysed. It was found that in the normal group, in patients aged 7-10, TT-PCL distance (p = 0.006), TTL (p = 0.007) and TT-PCL/PTW (p < 0.001) were significantly larger in females than in males. A significant sex effect was also detected on TT-TG/TW (p = 0.014). TT-TG distance, TT-PCL distance, TTL and TT-PCL/PTW (in male patients) approached an established normal adult value of 12.3 mm, 20.9 mm, 0.64 and 0.28, respectively, with increasing age (p < 0.001). The AUC was greater for TT-TG/TDI and TT-TG/TW (p ≤ 0.01) and TT-TG/TDI outperformed TT-TG/TW in patients aged 15-18 (p = 0.004). CONCLUSIONS: Tubercle lateralisation increased with age and was affected by sex, with the exception of TT-TG distance and TT-TG/TDI. TT-TG/TDI is the optimal method of diagnosing a lateralized tibial tubercle in patients with RPD. These findings assist with the evaluation of tubercle lateralisation in that they provide a proper protocol for paediatric and adolescent populations with RPD; and thus, will help determine whether medial tubercle transfer should be included among the tailored surgical procedures considered for the treatment of patients with RPD. LEVEL OF EVIDENCE: Level III.

The Arabidopsis TETRATRICOPEPTIDE-REPEAT THIOREDOXIN-LIKE (TTL) family members are involved in root system formation via their interaction with cytoskeleton and cell wall remodeling.

Lateral roots (LR) are essential components of the plant edaphic interface; contributing to water and nutrient uptake, biotic and abiotic interactions, stress survival, and plant anchorage. We have identified the TETRATRICOPEPTIDE-REPEAT THIOREDOXIN-LIKE 3 (TTL3) gene as being related to LR emergence and later development. Loss of function of TTL3 leads to a reduced number of emerged LR due to delayed development of lateral root primordia (LRP). This trait is further enhanced in the triple mutant ttl1ttl3ttl4. TTL3 interacts with microtubules and endomembranes, and is known to participate in the brassinosteroid (BR) signaling pathway. Both ttl3 and ttl1ttl3ttl4 mutants are less sensitive to BR treatment in terms of LR formation and primary root growth. The ability of TTL3 to modulate biophysical properties of the cell wall was established under restrictive conditions of hyperosmotic stress and loss of root growth recovery, which was enhanced in ttl1ttl3ttl4. Timing and spatial distribution of TTL3 expression is consistent with its role in development of LRP before their emergence and subsequent growth of LR. TTL3 emerged as a component of the root system morphogenesis regulatory network.

Biomechanical and clinical research of Isobar semi-rigid stabilization devices for lumbar degenerative diseases: a systematic review.

While lumbar spinal fusion using rigid rods is a prevalent surgical technique, it can lead to complications such as adjacent segment disease (ASDis). Dynamic stabilization devices serve to maintain physiological spinal motion and alleviate painful stress, yet they are accompanied by a substantial incidence of construct failure and subsequent reoperation. Compared to traditional rigid devices, Isobar TTL semi-rigid stabilization devices demonstrate equivalent stiffness and effective stabilization capabilities. Furthermore, when contrasted with dynamic stabilization techniques, semi-rigid stabilization offers improved load distribution, a broader range of motion within the fixed segment, and reduced mechanical failure rates. This paper will review and evaluate the clinical and biomechanical performance of Isobar TTL semi-rigid stabilization devices. A literature search using the PubMed, EMBASE, CNKI, Wanfang, VIP, and Cochrane Library databases identified studies that met the eligibility criteria. Twenty-eight clinical studies and nine biomechanical studies were included in this systematic review. The VAS, the ODI, and Japanese Orthopedic Association scoring improved significantly in most studies. UCLA grading scale, Pfirrmann grading, and modified Pfirrmann grading of the upper adjacent segments improved significantly in most studies. The occurrence rate of ASD was low. In biomechanical studies, Isobar TTL demonstrated a superior load sharing distribution, a larger fixed segment range of motion, and reduced stress at the rod-screw/screw-bone interfaces compared with titanium rods. While findings from mechanical studies provided promising results, the clinical studies exhibited low methodological quality. As a result, the available evidence does not possess sufficient strength to substantiate superior outcomes with Isobar semi-rigid system in comparison to titanium rods. To establish more conclusive conclusions, further investigations incorporating improved protocols, larger sample sizes, and extended follow-up durations are warranted.

Isobar hybrid dynamic stabilization with posterolateral fusion in mild and moderate lumbar degenerative disease.

OBJECTIVE: The aim of this study was to investigate the feasibility of using the Isobar TTL system and posterolateral fusion in a two-segment hybrid fixation approach, combined with spinal decompression, for treating mild and moderate lumbar degenerative disease. Specifically, we sought to evaluate the effectiveness of this approach for managing two-segment mild and moderate lumbar degenerative disease, and to determine whether it could provide a safe and reliable alternative to traditional surgical methods. METHODS: This retrospective study included 45 consecutive patients with two-level lumbar disc herniation or spinal stenosis, 24 of whom underwent the TTL system and posterolateral fusion combined (TTL group), and 21 of whom underwent posterolateral fusion alone (Rigid group). The surgical segment, admission diagnosis, operation time, and intraoperative bleeding were recorded separately for the two groups of patients. Imaging studies included pre- and postoperative radiography, magnetic resonance imaging, and computed tomography. The clinical outcomes were measured by Oswestry Disability Index (ODI) scores, and a visual analogue scale (VAS) for back and leg pain. RESULTS: All patients completed the surgery successfully with a mean follow-up of 56.09 months. The operative time and intraoperative bleeding were lower in the TTL group than in the Rigid group (p < 0.05). All patients showed significant improvements in clinical outcomes, including VAS for back and leg pain, and ODI scores (p < 0.05). ODI scores, the TTL group was better than the Rigid group at 1 year after surgery and at the final follow-up (p < 0.05). Postoperative surgical segment range of motion (ROM) decreased in both groups (p < 0.05). The postoperative ROM of the upper adjacent segment increased in both groups and was significantly higher in both groups at the last follow-up compared with the preoperative period (p < 0.05), and the superior adjacent segment rom of the TTL group was lower than the Rigid group (p < 0.05). The modified Pfrrmann classification of the superior adjacent segment was significantly increased in both groups at the last follow-up (p < 0.05). And in the TTL group, ROM, DH, and modified Pfrrmann grading of dynamic segment outperformed fusion segments. According to the UCLA classification, the incidence of adjacent segment degeneration (ASD) was 4.2% in the TTL group and 23.8% in the Rigid group, and the incidence of ASD was lower in the TTL group than in the Rigid group (P < 0.05). CONCLUSION: The Isobar TTL System was utilized in two-level lumbar hybrid surgery, resulting in no evident indications of lumbar instability being detected on X-rays captured at a minimum of 4 years after the operation, while retaining partial range of motion of the surgical segment. The general clinical efficacy is equivalent to titanium rod fusion surgery, presenting an alternative treatment for individuals with mild and moderate lumbar degenerative disease.

In COM we trust: Feasibility of USB-based event marking.

Modern experimental research often relies on the synchronization of different events prior to data analysis. One way of achieving synchronization involves marking distinct events with electrical pulses (event markers or "TTL pulses"), which are continuously recorded with research hardware, and can later be temporally aligned. Traditionally, this event marking was often performed using the parallel port in standard personal computers. However, the parallel port is disappearing from the landscape of computer hardware, being replaced by a serial (COM) port, namely the USB port. To find an adequate replacement for the parallel port, we evaluated four microcontroller units (MCUs) and the LabJack U3, an often-used USB data acquisition device, in terms of their latency and jitter for sending event markers in a simulated experiment on both Windows and Linux. Our results show that all four MCUs were comparable to the parallel port in terms of both latency and jitter, and consistently achieved latencies under 1 ms. With some caveats, the LabJack U3 can also achieve comparable latencies. In addition to the collected data, we share extensive documentation on how to build and use MCUs for event marking, including code examples. MCUs are a cost-effective, flexible, and performant replacement for the disappearing parallel port, enabling event marking and synchronization of data streams.

Analysis of clinical effect and radiographic outcomes of Isobar TTL system for two-segment lumbar degenerative disease: a retrospective study.

BACKGROUND: Nonfusion fixation is an effective way to treat lumbar degeneration. In the present study, we analyzed the clinical effects and radiographic outcomes of the Isobar TTL system used to treat two-segment lumbar degenerative disease. METHOD: Forty-one patients diagnosed with two-segment lumbar degenerative disease underwent surgical implantation of the Isobar TTL dynamic stabilization system (n = 20) or a rigid system (n = 21) from January 2013 to June 2017. The mean follow-up time was 23.6 (range 15-37) months. Clinical results were evaluated with the Oswestry Disability Index (ODI), modified Macnab criteria, and the visual analog score (VAS). Radiographic evaluations included the height of the intervertebral space and the range of motion (ROM) of the treated and adjacent segments. The intervertebral disc signal was classified using the modified Pfirrmann grading system and the University of California at Los Angeles (UCLA) system. RESULTS: The clinical results, including the ODI and VAS, showed that there was significant improvement in the two groups after implantation and that the difference between the two groups was not significant. In addition, the clinical efficacy indicated by the modified Macnab criteria for the two groups was similar. Radiological outcomes included the height of the intervertebral space, lumbar mobility, and intervertebral disc signal. The height of the intervertebral space of the upper adjacent segment L2/3 in the rigid group was significantly lower than that in the Isobar TTL group at the last follow-up. Furthermore, the number of ROMs of the fixed-segment L3/4 in the Isobar TTL group was significantly less than that before implantation, suggesting that the fixed-segment ROMs in the Isobar TTL group were limited. In addition, the ROM of the upper adjacent segment L2/3 in the last follow-up of the rigid group increased significantly, while that of the Isobar TTL group did not change after implantation. Finally, the incidence of adjacent-segment degeneration (ASD) was significantly greater in the rigid group than in the Isobar TTL group according to the UCLA system. CONCLUSION: The Isobar TTL system can be clinically effective for treating two-segment lumbar degenerative disease. Compared with rigid fixation, the Isobar TTL system yielded better radiographic outcomes and maintained the mobility of the treated segments with less impact on the proximal adjacent segment.

Tachypleus tridentatus Lectin Enhances Oncolytic Vaccinia Virus Replication to Suppress In Vivo Hepatocellular Carcinoma Growth.

Lectins play diverse roles in physiological processes as biological recognition molecules. In this report, a gene encoding Tachypleus tridentatus Lectin (TTL) was inserted into an oncolytic vaccinia virus (oncoVV) vector to form oncoVV-TTL, which showed significant antitumor activity in a hepatocellular carcinoma mouse model. Furthermore, TTL enhanced oncoVV replication through suppressing antiviral factors expression such as interferon-inducible protein 16 (IFI16), mitochondrial antiviral signaling protein (MAVS) and interferon-beta (IFN-ß). Further investigations revealed that oncoVV-TTL replication was highly dependent on ERK activity. This study might provide insights into a novel way of the utilization of TTL in oncolytic viral therapies.

The GSK3-MAP1B pathway controls neurite branching and microtubule dynamics.

The microtubule-associated protein MAP1B plays a key role in axon regeneration. We investigated the role of GSK3-mediated MAP1B phosphorylation in local fine-tuning of neurite branching and the underlying microtubule (MT) dynamics. In wildtype adult dorsal root ganglia (DRG) neurons, MAP1B phosphorylation is locally reduced at branching points, and branching dynamics from growth cones and distal neurite shafts is increased upon GSK3 inhibition. While map1b-/- neurites, that display increased branching, are not affected by GSK3 inhibition, transfection of map1b-/- neurons with full-length map1b-cDNA restores the wildtype branching phenotype, demonstrating that MAP1B is a key effector downstream of GSK3. Experiments in mutant mice lacking tyrosinated MTs indicate a preferential association of phospho-MAP1B with tyrosinated MTs. Interestingly, inhibition of GSK3-mediated MAP1B phosphorylation in map1b-cDNA-transfected fibroblasts protects both tyrosinated and acetylated MTs from nocodazole-induced depolymerization, while detyrosinated MTs are less abundant in the presence of MAP1B. Our data thus provide new insight into the molecular link between GSK3, MAP1B, neurite branching and MT stability regulation. We suggest that, at branching points, MAP1B undergoes a fine regulation of both its phosphorylation and sub-cellular amounts, in order to modulate the local balance between acetylated, detyrosinated, and tyrosinated microtubule pools.

Tubulin-tyrosine Ligase (TTL)-mediated Increase in Tyrosinated α-Tubulin in Injured Axons Is Required for Retrograde Injury Signaling and Axon Regeneration.

Injured peripheral neurons successfully activate a pro-regenerative program to enable axon regeneration and functional recovery. The microtubule-dependent retrograde transport of injury signals from the lesion site in the axon back to the cell soma stimulates the increased growth capacity of injured neurons. However, the mechanisms initiating this retrograde transport remain poorly understood. Here we show that tubulin-tyrosine ligase (TTL) is required to increase the levels of tyrosinated α-tubulin at the axon injury site and plays an important role in injury signaling. Preventing the injury-induced increase in tyrosinated α-tubulin by knocking down TTL impairs retrograde organelle transport and delays activation of the pro-regenerative transcription factor c-Jun. In the absence of TTL, axon regeneration is reduced severely. We propose a model in which TTL increases the levels of tyrosinated α-tubulin locally at the injury site to facilitate the retrograde transport of injury signals that are required to activate a pro-regenerative program.

Current Status: Site-Specific Antibody Drug Conjugates.

Antibody drug conjugates (ADCs), a promising class of cancer biopharmaceuticals, combine the specificity of therapeutic antibodies with the pharmacological potency of chemical, cytotoxic drugs. Ever since the first ADCs on the market, a plethora of novel ADC technologies has emerged, covering as diverse aspects as antibody engineering, chemical linker optimization and novel conjugation strategies, together aiming at constantly widening the therapeutic window for ADCs. This review primarily focuses on novel chemical and biotechnological strategies for the site-directed attachment of drugs that are currently validated for 2nd generation ADCs to promote conjugate homogeneity and overall stability.

A novel nematode effector suppresses plant immunity by activating host reactive oxygen species-scavenging system.

Evidence is emerging that plant-parasitic nematodes can secrete effectors to interfere with the host immune response, but it remains unknown how these effectors can conquer host immune responses. Here, we depict a novel effector, MjTTL5, that could suppress plant immune response. Immunolocalization and transcriptional analyses showed that MjTTL5 is expressed specifically within the subventral gland of Meloidogyne javanica and up-regulated in the early parasitic stage of the nematode. Transgenic Arabidopsis lines expressing MjTTL5 were significantly more susceptible to M. javanica infection than wild-type plants, and vice versa, in planta silencing of MjTTL5 substantially increased plant resistance to M. javanica. Yeast two-hybrid, coimmunoprecipitation and bimolecular fluorescent complementation assays showed that MjTTL5 interacts specifically with Arabidopsis ferredoxin : thioredoxin reductase catalytic subunit (AtFTRc), a key component of host antioxidant system. The expression of AtFTRc is induced by the infection of M. javanica. Interaction between AtFTRc and MjTTL could drastically increase host reactive oxygen species-scavenging activity, and result in suppression of plant basal defenses and attenuation of host resistance to the nematode infection. Our results demonstrate that the host ferredoxin : thioredoxin system can be exploited cunningly by M. javanica, revealing a novel mechanism utilized by plant-parasitic nematodes to subjugate plant innate immunity and thereby promoting parasitism.

Range aware message drop policy for delay tolerant networks.

In delay tolerant networks (DTNs) the messages are often not delivered to the destination due to a lack of end-to-end connectivity. In such cases, the messages are stored in the buffer for a long time and are transmitted when the nodes come into the range of each other. The buffer size of each node has a limited capacity, and it cannot accommodate the new incoming message when the buffer memory is full, and as a result network congestion occurs. This leads to a low delivery probability and thus increases the overhead ratio. In this research work, a new buffer management scheme called Range Aware Drop (RAD) is proposed which considers metrics such as message size and time to live (TTL). RAD utilizes TTL as an important metric and as a result, reduces the unnecessary message drop. Simulation results reveal that RAD performs significantly better than drop oldest (DOA) and size aware drop (SAD) in terms of delivery probability and overhead ratio. The obtained results also revealed that the hop-count average of SAD is 3.9 and DOA is 3.4 while the hop-count average of RAD is just 1.7. Also, the message drop ratio of the RAD is 36.2% while SAD and DOA have message drop ratios of 73.3% and 84.9% respectively.

Long-term Outcome of Isobar TTL System for the Treatment of Lumbar Degenerative Disc Diseases.

OBJECTIVE: The Isobar TTL dynamic fixation system has demonstrated favorable outcomes in the short-term treatment of lumbar degenerative disc diseases (LDDs). However, there is a paucity of extensive research on the long-term effects of this system on LDDs. This study aimed to evaluate the long-term clinical and radiological outcomes of patients with LDDs who underwent treatment utilizing the Isobar TTL dynamic fixation system. METHODS: The study analyzed the outcomes of 40 patients with LDDs who underwent posterior lumbar decompression and received single-segment Isobar TTL dynamic internal fixation at our hospital between June 2010 and December 2016. The evaluation of clinical therapeutic effect involved assessing postoperative pain levels using the visual analogue scale (VAS) and Oswestry disability index (ODI), both before surgery, 3 months after, and the final follow-up. To determine the preservation of functional motion in dynamically stable segments, we measured the range of motion (ROM) and disc height of stabilized and adjacent segments preoperatively and during the final follow-up. Additionally, we investigated the occurrence of adjacent segment degeneration (ASD). RESULTS: Forty patients were evaluated, with an average age of 44.65 years and an average follow-up period of 79.37 months. Fourteen patients belonged to the spondylolisthesis group, while the remaining 26 were categorized under the stenosis or herniated disc group. The preoperative ROM of the stabilized segment exhibited a significant reduction from 8.15° ± 2.77° to 5.00° ± 1.82° at the final follow-up (p < 0.001). In contrast, there was a slight elevation in the ROM of the adjacent segment during the final follow-up, increasing from 7.68° ± 2.25° before surgery to 9.36° ± 1.98° (p < 0.001). The intervertebral space height (IH) in the stabilized segment exhibited a significant increase from 10.56 ± 1.99 mm before surgery to 11.39 ± 1.90 mm at the one-week postoperative follow-up (p < 0.001). Conversely, there was a notable decrease in the IH of the adjacent segment from 11.09 ± 1.82 mm preoperatively to 10.86 ± 1.79 mm at the one-week follow-up after surgery (p < 0.001). The incidence of ASD was 15% (6/40) after an average follow-up period of 79.37 months, with a rate of 15.38% (4/26) in the stenosis or herniated disc group and 14.29% (2/14) in the spondylolisthesis group; however, no statistically significant difference was observed in the occurrence of ASD among these groups (p > 0.05). CONCLUSION: The Isobar TTL dynamic fixation system is an effective treatment for LDDs, improving pain relief, quality of life (QoL) and maintaining stabilized segmental motion. It has demonstrated excellent long-term clinical and radiographic results.

Clinical Outcomes of Isobar TTL System with Isthmic Bone Grafting and Pedicle Screw-Vertebral Plate Hook with Direct Repair of Defect for Lumbar Spondylolysis: A Matched-Pair Case Control Study.

OBJECTIVE: Although direct isthmic repair, such as PSVPH, did not affect the mobility of the fixed segment and adjacent segment, it has a relatively low rate of isthmic fusion compared with conventional fusion. The Isobar TTL dynamic internal fixation system has been widely used in clinical practice and has achieved satisfactory clinical results. However, the use of the Isobar TTL system in combination with direct isthmic repair for lumbar spondylolysis has rarely been reported. The aim of this study was to compare the clinical and radiologic outcomes between patients who underwent Isobar TTL system and PSVPH with direct repair of defect for lumbar spondylolysis. METHODS: Stepwise propensity score matching (PSM) for age and sex were performed to keep comparable clinical data between groups in this retrospective and matched-pair case control study. A total of 50 patients diagnosed with lumbar spondylolysis underwent surgical implantation of the Isobar TTL group (n = 25) or PSVPH group (n = 25) from June 2009 to June 2016. Clinical outcomes were assessed using the Oswestry disability index (ODI), and visual analog score (VAS). Radiographic evaluations included range of motion (ROM) and the disc heights of stabilized segment and adjacent segment, adjacent segment degeneration (ASD) and bony fusion. Three-dimensional reconstruction of lumbar CT scan was obtained to evaluate bone fusion of the isthmic at final follow-up. The independent Student's t test and chi-square test were applied to compare the differences between groups. RESULTS: A total of 25 patients from TTL group were matched to 25 patients in PSVPH group for age, sex, body mass index (BMI), defect side, spondylolisthesis meyerding, and follow-up duration. The intervertebral space height (IH) of stabilized segment at postoperative 1 week and final follow-up in the TTL group was higher than those in the PSVPH group, respectively (P = 0.030; P = 0.013). The ROM of stabilized segment at final follow-up in the TTL group was significantly lower than that in the PSVPH group (P < 0.001). The bony fusion rate at the final follow-up was 88.0% (22/25 cages) in the TTL group and 80.0% (20/25 cages) in the PSVPH group. The ODI score at final follow-up in the TTL group was significantly lower than that in the PSVPH group (P = 0.007). CONCLUSION: Overall, our data suggest that the Isobar TTL system outcomes are comparable to those in the PSVPH, with a similar high bony fusion rate as PSVPH, especially its wider indications as a new surgery.

Immune and Reproductive Biomarkers in Female Sea Urchins Paracentrotus lividus under Heat Stress.

The functioning of the immune and reproductive systems is crucial for the fitness and survival of species and is strongly influenced by the environment. To evaluate the effects of short-term heat stress (HS) on these systems, confirming and deepening previous studies, female sea urchin Paracentrotus lividus were exposed for 7 days to 17 °C, 23 and 28 °C. Several biomarkers were detected such as the ferric reducing power (FRAP), ABTS-based total antioxidant capacity (TAC-ABTS), nitric oxide metabolites (NOx), total thiol levels (TTL), myeloperoxidase (MPO) and protease (PA) activities in the coelomic fluid (CF) and mitochondrial membrane potential (MMP), H2O2 content and intracellular pH (pHi) in eggs and coelomocytes, in which TAC-ABTS and reactive nitrogen species (RNS) were also analyzed. In the sea urchins exposed to HS, CF analysis showed a decrease in FRAP levels and an increase in TAC-ABTS, TTL, MPO and PA levels; in coelomocytes, RNS, MMP and H2O2 content increased, whereas pHi decreased; in eggs, increases in MMP, H2O2 content and pHi were found. In conclusion, short-term HS leads to changes in five out of the six CF biomarkers analyzed and functional alterations in the cells involved in either reproductive or immune activities.

MicroFPGA: An affordable FPGA platform for microscope control.

Modern microscopy relies increasingly on microscope automation to improve throughput, ensure reproducibility or observe rare events. Automation requires computer control of the important elements of the microscope. Furthermore, optical elements that are usually fixed or manually movable can be placed on electronically-controllable elements. In most cases, a central electronics board is necessary to generate the control signals they require and to communicate with the computer. For such tasks, Arduino microcontrollers are widely used due to their low cost and programming entry barrier. However, they are limiting in their performance for applications that require high-speed or multiple parallel processes. Field programmable gate arrays (FPGA) are the perfect technology for high-speed microscope control, as they are capable of processing signals in parallel and with high temporal precision. While plummeting prices made the technology available to consumers, a major hurdle remaining is the complex languages used to configure them. In this work, we used an affordable FPGA, delivered with an open-source and friendly-to-use programming language, to create a versatile microscope control platform called MicroFPGA. It is capable of synchronously triggering cameras and multiple lasers following complex patterns, as well as generating various signals used to control microscope elements such as filter wheels, servomotor stages, flip-mirrors, laser power or acousto-optic modulators. MicroFPGA is open-source and we provide online Micro-Manager, Java, Python and LabVIEW libraries, together with blueprints and tutorials.

Comparison of Tibial Tubercle Landmark Technique and Range of Motion Technique in Primary Total Knee Arthroplasty: A Retrospective Cohort Study.

OBJECTIVE: There is not a standard for rotational alignment of the tibial component in total knee arthroplasty (TKA). For now, the most commonly methods are tibial-tubercle -landmark technique (TTL) and range-of-motion technique (ROM). The study is aimed to compare clinical outcomes and radiographic data of patients who undergone primary TKA with TTL or ROM technique. METHODS: This single-surgeon retrospective cohort study includes 60 patients with TTL technique and 60 with ROM technique from December 2017 to January 2019. All patients were evaluated clinically using Hospital for Special Surgery Knee Score (HSS), Feller patellar score, visual analogue scale (VAS) and maximum knee flexion and extension angle before and after surgery at both 6 months and 12 months postoperatively. Radiographic data contain hip-knee-ankle angle (HKA), mechanical lateral distal femoral angle (mLDFA), mechanical medial proximal tibial angle (mMPTA), posterior slope angle (PSA) on pre and postoperative X-ray and rotation angle of femoral component (relative to surgical trans-epicondylar axis) and tibial component (relative to surgical trans-epicondylar axis, tibial posterior condylar line and Akagi') on postoperative computed tomography (CT) scan. Clinical outcomes and radiological data were compared between the two groups. RESULTS: One hundred twenty patients (120 knees) were enrolled in this study, including 38 males and 82 females, aged from 58 to 78, with an average of 65.7 years. There was no significant difference in demographics and preoperative X-ray data between the two groups (P > 0. 05). Clinical scores of the TTL group were better than those in the ROM group at 6 and 12 months after surgery, when comparing HSS (83.57 ± 5.00 vs 75.90 ± 4.89, F = 59.004, P < 0.001; 90.53 ± 4.31 vs 82.83 ± 4.98, F = 54.509, P < 0.001), Feller patellar score (21.43 ± 2.54 vs 19.10 ± 2.52, F = 14.864, P = 0.001; 26.27 ± 1.98 vs 23.20 ± 2.31, F = 42.204, P < 0.001) and VAS (3.70 ± 0.62 vs 4.38 ± 0.92, F = 14.508, P = 0.001; 2.10 ± 0.90 vs 2.79 ± 0.80, F = 11.554, P = 0.002). But there was no significant difference in the flexion and extension angle between the two groups. In imaging evaluation, no statistical difference was found in pre- and postoperative HKA, mLDFA, mMPTA and PSA. Rotational angles of tibial component only did relative to Akagi' have statistical difference in two groups (2.33 ± 4.3 vs 4.41 ± 3.2, t = 2.143, P < 0.05) (Positive value represented external rotation). CONCLUSION: The results of our study showed that both methods were reliable, and TTL technique provided better clinical scores and larger external angle of tibial component, compared to ROM technique.

Residue-Specific Incorporation of the Non-Canonical Amino Acid Norleucine Improves Lipase Activity on Synthetic Polyesters.

Environmentally friendly functionalization and recycling processes for synthetic polymers have recently gained momentum, and enzymes play a central role in these procedures. However, natural enzymes must be engineered to accept synthetic polymers as substrates. To enhance the activity on synthetic polyesters, the canonical amino acid methionine in Thermoanaerobacter thermohydrosulfuricus lipase (TTL) was exchanged by the residue-specific incorporation method for the more hydrophobic non-canonical norleucine (Nle). Strutural modelling of TTL revealed that residues Met-114 and Met-142 are in close vicinity of the active site and their replacement by the norleucine could modulate the catalytic activity of the enzyme. Indeed, hydrolysis of the polyethylene terephthalate model substrate by the Nle variant resulted in significantly higher amounts of release products than the Met variant. A similar trend was observed for an ionic phthalic polyester containing a short alkyl diol (C5). Interestingly, a 50% increased activity was found for TTL [Nle] towards ionic phthalic polyesters containing different ether diols compared to the parent enzyme TTL [Met]. These findings clearly demonstrate the high potential of non-canonical amino acids for enzyme engineering.

Manipulation of the Tubulin Code Alters Directional Cell Migration and Ciliogenesis.

Conjunction of epithelial cells into monolayer sheets implies the ability to migrate and to undergo apicobasal polarization. Both processes comprise reorganization of cytoskeletal elements and rearrangements of structural protein interactions. We modulated expression of tubulin tyrosin ligase (TTL), the enzyme that adds tyrosine to the carboxy terminus of detyrosinated α-tubulin, to study the role of tubulin detyrosination/-tyrosination in the orientation of cell motility and in epithelial morphogenesis. Oriented cell migration and the organization of focal adhesions significantly lose directionality with diminishing amounts of microtubules enriched in detyrosinated tubulin. On the other hand, increasing quantities of detyrosinated tubulin results in faster plus end elongation of microtubules in migrating and in polarized epithelial cells. These plus ends are decorated by the plus end binding protein 1 (EB1), which mediates interaction between microtubules enriched in detyrosinated tubulin and the integrin-ILK complex at focal adhesions. EB1 accumulates at the apical cell pole at the base of the primary cilium following apicobasal polarization. Polarized cells almost devoid of detyrosinated tubulin form stunted primary cilia and multiluminal cysts in 3D-matrices. We conclude that the balance between detyrosinated and tyrosinated tubulin alters microtubule dynamics, affects the orientation of focal adhesions and determines the organization of primary cilia on epithelial cells.