Dropped Head Syndrome Attenuation by Hybrid Assistive Limb: A Preliminary Study of Three Cases on Cervical Alignment during Walking.

Background and Objectives: Dropped head syndrome (DHS) is characterized by apparent neck extensor muscle weakness and difficulty in extending the neck to raise the head against gravity. DHS affects forward vision and eating behavior, and hence impairs quality of life. However, standardized treatment of DHS has not yet been established. The purpose of this preliminary study is to seek for a possibility of effective non-surgical, conservative treatment for DHS, by applying a robotic treatment. Materials and Methods: A wearable exoskeleton type robot suit hybrid assistive limb (HAL) was applied to three patients with DHS. A course of HAL treatment included 10 sessions of gait training using HAL. One session lasted about an hour. Case 1 completed the course twice, the first time in two weeks (one session per day) and second time in 10 months (one session per month). Case 2 and Case 3 completed the course once in 10 weeks (one session per week) and in 6 months (one session per 2.5 weeks), respectively. Immediate and lasting effects of HAL on the reduction of cervical sagittal vertical alignment (SVA) during gait was evaluated using a motion capture system. Results: Case 1 showed improvement of cervical alignment during gait after the HAL courses of both different frequencies. Case 2 did not show improvement of cervical alignment during gait. Case 3 showed improvement of cervical kyphosis but not of cervical sagittal alignment during gait. Conclusions: The results of the preliminary study suggest that gait training using HAL may be an effective option of conservative treatment for a part of DHS patients. They also suggest that a lack of immediate effects on the cervical alignment and a lack of ability to perform compensatory trunk motion may indicate a non-responding patient. Generalization of the results requires further research with more cases.

HAL2 overexpression induces iron acquisition in bdf1Δ cells and enhances their salt resistance.

The yeast Saccharomyces cerevisiae is capable of responding to various environmental stresses, such as salt stress. Such responses require a complex network and adjustment of the gene expression network. The goal of this study is to further understand the molecular mechanism of salt stress response in yeast, especially the molecular mechanism related to genes BDF1 and HAL2. The Bromodomain Factor 1 (Bdf1p) is a transcriptional regulator, which is part of the basal transcription factor TFIID. Cells lacking Bdf1p are salt sensitive with an abnormal mitochondrial function. We previously reported that the overexpression of HAL2 or deletion of HDA1 lowers the salt sensitivity of bdf1Δ. To better understand the mechanism behind the HAL2-related response to salt stress, we compared three global transcriptional profiles (bdf1Δ vs WT, bdf1Δ + HAL2 vs bdf1Δ, and bdf1Δhda1Δ vs bdf1Δ) in response to salt stress using DNA microarrays. Our results reveal that genes for iron acquisition and cellular and mitochondrial remodeling are induced by HAL2. Overexpression of HAL2 decreases the concentration of nitric oxide. Mitochondrial iron-sulfur cluster (ISC) assembly also decreases in bdf1Δ + HAL2. These changes are similar to the changes of transcriptional profiles induced by iron starvation. Taken together, our data suggest that mitochondrial functions and iron homeostasis play an important role in bdf1Δ-induced salt sensitivity and salt stress response in yeast.

Lipopolysaccharide induces amyloid formation of antimicrobial peptide HAL-2.

Lipopolysaccharide (LPS), the important component of the outer membrane of Gram-negative bacteria, contributes to the integrity of the outer membrane and protects the cell against bactericidal agents, including antimicrobial peptides. However, the mechanisms of interaction between antimicrobial peptides and LPS are not clearly understood. Halictines-2 (HAL-2), one of the novel antimicrobial peptides, was isolated from the venom of the eusocial bee Halictus sexcinctus. HAL-2 has exhibited potent antimicrobial activity against Gram-positive and Gram-negative bacteria and even against cancer cells. Here, we studied the interactions between HAL-2 and LPS to elucidate the antibacterial mechanism of HAL-2 in vitro. Our results show that HAL-2 adopts a significant degree of ß-strand structure in the presence of LPS. LPS is capable of inducing HAL-2 amyloid formation, which may play a vital role in its antimicrobial activity.

Yeast 3',5'-bisphosphate nucleotidase: an affinity tag for protein purification.

Affinity chromatography is one of the most popular methods for protein purification. Each tag method has its advantages and disadvantages, and combination of different tags and developing of new tags had been proposed and performed. Yeast 3',5'-bisphosphate nucleotidase, also known as HAL2, hydrolyzes 3'-phosphoadenosine 5'-phosphate (PAP) with submicromolar Km, which indicated the tight interactions between HAL2 and PAP. In order to explore the feasibility of HAL2 as a protein purification affinity tag, HAL2 was further characterized with PAP as substrate. Results demonstrated that KmPAP and kcatPAP were ∼0.3µM and ∼11s(-)(1), respectively. Kd for PAP was 0.008µM in the presence of Ca(2+). pH was also found to affect interactions between HAL2 and PAP, with tightest binding (Kd∼8nM) at pH 7.5 and 8. The purification protocol was rationally designed based on nanomolar affinity to PAP agarose in the presence of Ca(2+), which could satisfy the metal requirement for PAP binding, prevent hydrolysis of immobilized PAP and could be chelated by ethylene glycol tetraacetic acid (EGTA) for elution. A series of expression vectors were further constructed and Escherichia coli adenosine 5'-phosphosulfate kinase (APSK) was prokaryotically expressed, purified and characterized. Ready to use expression vector with eight commonly used restriction enzyme recognition sites in multiple cloning site was subsequently constructed. By comparing with current popular tags, HAL2 was found to be an efficient and economical tag for prokaryotic protein expression and purification.

Adaptation of extremely halotolerant black yeast Hortaea werneckii to increased osmolarity: a molecular perspective at a glance.

Halophilic adaptations have been studied almost exclusively on prokaryotic microorganisms. Discovery of the black yeast Hortaea werneckii as the dominant fungal species in hypersaline waters enabled the introduction of a new model organism to study the mechanisms of salt tolerance in eukaryotes. Its strategies of cellular osmotic adaptations on the physiological and molecular level revealed novel, intricate mechanisms to combat fluctuating salinity. H. werneckii is an extremely halotolerant eukaryotic microorganism and thus a promising source of transgenes for osmotolerance improvement of industrially important yeasts, as well as in crops.