Deadenylation kinetics of mixed poly(A) tails at single-nucleotide resolution.

Shortening of messenger RNA poly(A) tails, or deadenylation, is a rate-limiting step in mRNA decay and is highly regulated during gene expression. The incorporation of non-adenosines in poly(A) tails, or 'mixed tailing', has been observed in vertebrates and viruses. Here, to quantitate the effect of mixed tails, we mathematically modeled deadenylation reactions at single-nucleotide resolution using an in vitro deadenylation system reconstituted with the complete human CCR4-NOT complex. Applying this model, we assessed the disrupting impact of single guanosine, uridine or cytosine to be equivalent to approximately 6, 8 or 11 adenosines, respectively. CCR4-NOT stalls at the 0, -1 and -2 positions relative to the non-adenosine residue. CAF1 and CCR4 enzyme subunits commonly prefer adenosine but exhibit distinct sequence selectivities and stalling positions. Our study provides an analytical framework to monitor deadenylation and reveals the molecular basis of tail sequence-dependent regulation of mRNA stability.

Effects of SKCPT on Osteoarthritis in Beagle Meniscectomy and Cranial Cruciate Ligament Transection Models.

Osteoarthritis (OA) affects >500 million people globally, and this number is expected to increase. OA management primarily focuses on symptom alleviation, using non-steroidal anti-inflammatory drugs, including Celecoxib. However, such medication has serious side effects, emphasizing the need for disease-specific treatment. The meniscectomy and cranial cruciate ligament transection (CCLx)-treated beagle dog was used to investigate the efficacy of a modified-release formulation of SKI306X (SKCPT) from Clematis mandshurica, Prunella vulgaris, and Trichosanthes kirilowii in managing arthritis. SKCPT's anti-inflammatory and analgesic properties have been assessed via stifle circumference, gait, incapacitance, histopathology, and ELISA tests. The different SKCPT concentrations and formulations also affected the outcome. SKCPT improved the gait, histopathological, and ELISA OA assessment parameters compared to the control group. Pro-inflammatory cytokines and matrix metalloproteinases were significantly lower in the SKCPT-treated groups than in the control group. This study found that SKCPT reduces arthritic lesions and improves abnormal gait. The 300 mg modified-release formulation was more efficacious than others, suggesting a promising approach for managing OA symptoms and addressing disease pathogenesis. A high active ingredient level and a release pattern make this formulation effective for twice-daily arthritis treatment.

Telomeres reforged with non-telomeric sequences in mouse embryonic stem cells.

Telomeres are part of a highly refined system for maintaining the stability of linear chromosomes. Most telomeres rely on simple repetitive sequences and telomerase enzymes to protect chromosomal ends; however, in some species or telomerase-defective situations, an alternative lengthening of telomeres (ALT) mechanism is used. ALT mainly utilises recombination-based replication mechanisms and the constituents of ALT-based telomeres vary depending on models. Here we show that mouse telomeres can exploit non-telomeric, unique sequences in addition to telomeric repeats. We establish that a specific subtelomeric element, the mouse template for ALT (mTALT), is used for repairing telomeric DNA damage as well as for composing portions of telomeres in ALT-dependent mouse embryonic stem cells. Epigenomic and proteomic analyses before and after ALT activation reveal a high level of non-coding mTALT transcripts despite the heterochromatic nature of mTALT-based telomeres. After ALT activation, the increased HMGN1, a non-histone chromosomal protein, contributes to the maintenance of telomere stability by regulating telomeric transcription. These findings provide a molecular basis to study the evolution of new structures in telomeres.

Quantification of purified endogenous miRNAs with high sensitivity and specificity.

MicroRNAs (miRNAs) are short (19-24 nt) non-coding RNAs that suppress the expression of protein coding genes at the post-transcriptional level. Differential expression profiles of miRNAs across a range of diseases have emerged as powerful biomarkers, making a reliable yet rapid profiling technique for miRNAs potentially essential in clinics. Here, we report an amplification-free multi-color single-molecule imaging technique that can profile purified endogenous miRNAs with high sensitivity, specificity, and reliability. Compared to previously reported techniques, our technique can discriminate single base mismatches and single-nucleotide 3'-tailing with low false positive rates regardless of their positions on miRNA. By preloading probes in Thermus thermophilus Argonaute (TtAgo), miRNAs detection speed is accelerated by more than 20 times. Finally, by utilizing the well-conserved linearity between single-molecule spot numbers and the target miRNA concentrations, the absolute average copy numbers of endogenous miRNA species in a single cell can be estimated. Thus our technique, Ago-FISH (Argonaute-based Fluorescence In Situ Hybridization), provides a reliable way to accurately profile various endogenous miRNAs on a single miRNA sensing chip.

Accurate concentration control of mitochondria and nucleoids.

All cellular materials are partitioned between daughters at cell division, but by various mechanisms and with different accuracy. In the yeast Schizosaccharomyces pombe, the mitochondria are pushed to the cell poles by the spindle. We found that mitochondria spatially reequilibrate just before division, and that the mitochondrial volume and DNA-containing nucleoids instead segregate in proportion to the cytoplasm inherited by each daughter. However, nucleoid partitioning errors are suppressed by control at two levels: Mitochondrial volume is actively distributed throughout a cell, and nucleoids are spaced out in semiregular arrays within mitochondria. During the cell cycle, both mitochondria and nucleoids appear to be produced without feedback, creating a net control of fluctuations that is just accurate enough to avoid substantial growth defects.

Carbon monoxide-induced cardiomyopathy.

BACKGROUND: Previous reports demonstrated mechanisms of cardiac toxicity in acute carbon monoxide (CO) poisoning. Still, none established CO-induced cardiomyopathy (CMP) as a clinical entity. The aim of this study is to investigate CO-induced CMP in patients with acute CO poisoning in terms of its epidemiology, clinical characteristics, and prognosis. METHODS AND RESULTS: A retrospective study was conducted on consecutive patients who were diagnosed with acute CO poisoning at the emergency department of Ajou University Hospital during the period of 62 month. Six hundred and twenty-six patients were diagnosed with acute CO poisoning. During the initial echocardiography, 19 patients were abnormal: (1) global hypokinesia/akinesia (n=7), (2) regional wall hypokinesia/akinesia [n=12; takotsubo type (n=6), reverse takotsubo type (n=2), non-specific type (n=4)]. The ejection fraction (EF) was 36.3±13.5% (from 15% to 55%) and less than 45% for 14 patients. In the follow-up echocardiography performed within 12 days after the initial performance, most patients were found to have cardiac wall motion abnormalities, and their EF had returned to normal (ie, EF ≥50%). CONCLUSIONS: CO-induced CMP was identified in 3.04% (n=19) of all patients (n=626). It might not be too critical in acute clinical courses of acute CO poisoning because the prognosis seems favorable. Considering the common factors between CO-induced CMP and takotsubo CMP, myocardial stunning subject to a catecholamine surge most likely plays a central role in the development of CO-induced CMP.

Injuries associated with the 580 km university student grand voluntary road march: focus on foot injuries.

College student volunteers (n = 142) completed a 580 km road march for 21 consecutive days. Each volunteer carried a backpack that weighed 14.1 ± 1.4 kg on the average. We investigated the incidence and location of blisters associated with the road march using a foot map along with other injuries. Overall, 95.1% of the subjects (135 of 142) sustained one or more injuries. All injured subjects had foot blisters, and 18% had other foot injuries. The most common locations of blister development were the right 5th toe (61%) and the left 5th toe (57%). The little toes seem to have been subjected to the greatest friction and shearing forces. March-related injuries, excluding foot injuries, were ankle pain (12.7%), knee pain (12.7%) and Achilles tendon pain (7.7%). Six subjects (4.2%) needed extra medical treatment for more than 2 weeks prior to returning to their daily lives after completion of the march due to associated injuries. The present study observed a very high incidence rate of injuries (95.1%) associated with the 580 km university students grand road march. These injuries posed an obstacle against completion of the road march and against returning to daily life. Active preventive interventions such as physical therapy and customized reinforced shoes and education program are recommended for reducing incidence rate and severity of injuries.

The usefulness of the serum s100b protein for predicting delayed neurological sequelae in acute carbon monoxide poisoning.

CONTEXT: Some patients with acute carbon monoxide poisoning will experience delayed neurological sequelae. Several factors associated with delayed neurological sequelae have been reported, but these factors are unsatisfactory for the assessment of unconscious patients. OBJECTIVE: The aim of this study was to assess the usefulness of the serum S100B protein as a biochemical marker for predicting delayed neurological sequelae. MATERIALS AND METHODS: In this retrospective study, we evaluated the data for patients who visited an emergency medical center once during a period of 7 months. The enrollment criteria were the diagnosis of acute carbon monoxide poisoning and the measurement of the serum S100B level. A standardized extraction using medical records was performed. RESULT: A total of 71 patients were enrolled, and 10 patients (14.1%) presented delayed neurological sequelae. The delayed neurological sequelae group had a longer duration of carbon monoxide exposure, a longer duration of loss of consciousness, and a worse mental status (p-value < 0.001). In addition, the S100B protein levels were higher in the delayed neurological sequelae group (0.891 vs. 0.063, p-value < 0.001). Multiple logistic regression analysis showed that only the serum S100B protein level was independently associated with the development of delayed neurological sequelae (OR, 120.594; 95% CI, 4.194-3467.220), and a serum S100B protein level of more than 0.165 µg/L predicted the development of delayed neurological sequelae (sensitivity 90%, specificity 87%). DISCUSSION AND CONCLUSION: In the present study, the level of serum S100B protein was found to be useful for evaluating acute CO poisoning patients and was found to be an independent predictor of the development of DNS after acute CO poisoning.

Three-dimensional fluidic self-assembly by axis translation of two-dimensionally fabricated microcomponents in railed microfluidics.

A method for high-throughput 3D self-assembly of 2D photopatterned microstructures using railed microfluidics is presented. Vertical device patterning of heterogeneous materials requires high-level integration using conventional microelectromechanical system (MEMS) technology; however, 3D railed assembly enables easy and fast self-assembly via a fluidic axis-translation process and simple material exchange in microfluidic channels. Individually photopatterned 2D microstructures are axis-translated from in-plane to out-of-plane and fluidically self-assembled, guided by side-rails in microfluidic channels to form a 3D morphology. Since the structures are fabricated in fluidic environments, there are no fixed initial points on the channel substrate allowing fluidic horizontal stacking of erected 2D structures. The guiding mechanism of railed microfluidics enables efficient fluidic handling and deterministic 3D self-assembly of heterogeneous components such as electronic components or polymeric microstructures using only fluidic force.

The Effectiveness of Mobile Discharge Instruction Videos (MDIVs) in communicating discharge instructions to patients with lacerations or sprains.

OBJECTIVE: This study evaluates the effectiveness of mobile discharge instruction videos (MDIVs) in communicating discharge instructions to patients with lacerations or sprains. METHOD: A prospective controlled study was performed on patients with lacerations or sprains in a quaternary emergency center from April 1, 2008 to May 31, 2008. Upon discharge, patients were systematically allocated to receive printed discharge instructions (PDIs) or MDIVs. Within 48 hours of the patients' discharge, a standard questionnaire was provided via telephone to evaluate the patients' comprehension of, convenience rating for, and satisfaction with their given discharge instructions. RESULTS: Of the 645 patients with lacerations or sprains during the study period, a convenience sample of 161 patients was enrolled in the study; 77 in the PDIs group (the P group) and 84 in the MDIVs group (the M group). There were no statistically significant differences in the ages, genders, and levels of education of the subjects in the two groups. The mean of the correct answers on wound care in the questionnaire was 2.7 +/- 0.7 in the M group and 2.4 +/- 0.8 in the P group (P < 0.05). The convenience rating was 85.7% in the M group and 63.6% in the P group (P < 0.05). The rate of satisfaction was 90.5% in the M group and 90.9% in the P group (P > 0.05). CONCLUSION: The mobile discharge instruction video (MDIV) improved the communication of discharge instructions. Further studies will be needed to explore the actual compliance of patients to treatment regimens.