Dexamethasone reduces cisplatin-induced hair cell damage by inducing cisplatin resistance through metallothionein-2.

PURPOSE: Hair cell damage is a common side effect caused by the anticancer drug cisplatin (CDDP), which reduces patient quality of life. One CDDP resistance mechanism that occurs in recurrent cancers is heavy metal detoxification by metallothionein-2 (mt2). Here, we show that in zebrafish larvae, dexamethasone (DEX) reduces CDDP-induced hair cell damage by enhancing mt2 expression. METHODS: Transgenic zebrafish (cldn: gfp; atoh1: rfp) that express green and red fluorescent proteins in neuromasts and hair cells, respectively, were used. The zebrafish were pretreated with DEX at 52 h post-fertilization (hpf) for 8 h, followed by CDDP treatment for 12 h. The lateral line hair cells of CDDP-treated zebrafish at 72 hpf were observed by fluorescence microscopy. RESULTS: Reporting odds ratio (ROR) analysis using an adverse event database indicated an association between a decrease in CDDP-induced ototoxicity and DEX as an antiemetic treatment for cancer chemotherapy. Pretreatment with DEX protected 72 hpf zebrafish hair cells from CDDP-induced damage. The expression of mt2 mRNA was significantly increased by the combination of 10 µM DEX with CDDP. Gene editing of mt2 reversed the protective effect of DEX against CDDP-induced damage in hair cells. CONCLUSION: DEX protects hair cells from CDDP-induced damage through increased mt2 expression, which is a resistance mechanism for platinum-based anticancer drugs.

Interpersonal counseling versus active listening in the treatment of mild depression: a randomized controlled trial.

[Purpose] Healthcare workers, such as physical therapists, need to be equipped in dealing with patients' psychological problems. The three-session interpersonal counseling (three-session IPC) is a constructed counseling method that can be performed even by non-mental health professionals. This study examined the efficacy of the three-session IPC for treating depression. Immediate efficacy and efficacy up to 12 weeks post-intervention were examined. [Participants and Methods] In this randomized controlled trial of the two groups, one group (n=24) received the three-session IPC therapy (IPC group) while the other (n=24) received three sessions of active listening (active listening group). Depression was assessed using the Self-Rating Depression Scale (SDS) at baseline, post-intervention, and at 4, 8, and 12 weeks. [Results] There was a significant difference in total SDS scores between the IPC and active listening groups from baseline to 4 weeks after counseling, although no significant differences were observed at other time points. [Conclusion] The three-session IPC may be effective for 4 weeks after counseling. However, further studies are warranted in this regard.

Metastasis from the tumor interior and necrotic core formation are regulated by breast cancer-derived angiopoietin-like 7.

Necrosis in the tumor interior is a common feature of aggressive cancers that is associated with poor clinical prognosis and the development of metastasis. How the necrotic core promotes metastasis remains unclear. Here, we report that emergence of necrosis inside the tumor is correlated temporally with increased tumor dissemination in a rat breast cancer model and in human breast cancer patients. By performing spatially focused transcriptional profiling, we identified angiopoietin-like 7 (Angptl7) as a tumor-specific factor localized to the perinecrotic zone. Functional studies showed that Angptl7 loss normalizes central necrosis, perinecrotic dilated vessels, metastasis, and reduces circulating tumor cell counts to nearly zero. Mechanistically, Angptl7 promotes vascular permeability and supports vascular remodeling in the perinecrotic zone. Taken together, these findings show that breast tumors actively produce factors controlling central necrosis formation and metastatic dissemination from the tumor core.

Orchestration of Collective Migration and Metastasis by Tumor Cell Clusters.

Metastatic dissemination has lethal consequences for cancer patients. Accruing evidence supports the hypothesis that tumor cells can migrate and metastasize as clusters of cells while maintaining contacts with one another. Collective metastasis enables tumor cells to colonize secondary sites more efficiently, resist cell death, and evade the immune system. On the other hand, tumor cell clusters face unique challenges for dissemination particularly during systemic dissemination. Here, we review recent progress toward understanding how tumor cell clusters overcome these disadvantages as well as mechanisms they utilize to gain advantages throughout the metastatic process. We consider useful models for studying collective metastasis and reflect on how the study of collective metastasis suggests new opportunities for eradicating and preventing metastatic disease.

Sulfated Hyaluronan Binds to Heparanase and Blocks Its Enzymatic and Cellular Actions in Carcinoma Cells.

We examined whether sulfated hyaluronan exerts inhibitory effects on enzymatic and biological actions of heparanase, a sole endo-beta-glucuronidase implicated in cancer malignancy and inflammation. Degradation of heparan sulfate by human and mouse heparanase was inhibited by sulfated hyaluronan. In particular, high-sulfated hyaluronan modified with approximately 2.5 sulfate groups per disaccharide unit effectively inhibited the enzymatic activity at a lower concentration than heparin. Human and mouse heparanase bound to immobilized sulfated hyaluronan. Invasion of heparanase-positive colon-26 cells and 4T1 cells under 3D culture conditions was significantly suppressed in the presence of high-sulfated hyaluronan. Heparanase-induced release of CCL2 from colon-26 cells was suppressed in the presence of sulfated hyaluronan via blocking of cell surface binding and subsequent intracellular NF-κB-dependent signaling. The inhibitory effect of sulfated hyaluronan is likely due to competitive binding to the heparanase molecule, which antagonizes the heparanase-substrate interaction. Fragment molecular orbital calculation revealed a strong binding of sulfated hyaluronan tetrasaccharide to the heparanase molecule based on electrostatic interactions, particularly characterized by interactions of (-1)- and (-2)-positioned sulfated sugar residues with basic amino acid residues composing the heparin-binding domain-1 of heparanase. These results propose a relevance for sulfated hyaluronan in the blocking of heparanase-mediated enzymatic and cellular actions.

Depression Among University Students With Attention-Deficit Hyperactivity Disorder Symptoms: A Study of Interpersonal Counseling.

OBJECTIVE: University students with symptoms of attention-deficit hyperactivity disorder (ADHD) often experience depression. This study examined whether interpersonal counseling (IPC) could be an effective treatment for depression among students with ADHD symptoms. METHODS: Participants were assigned to either an IPC (N=5) or control (N=7) group. Depression was assessed by using the Self-Rating Depression Scale (SDS) at baseline, postintervention, and at 4-, 8-, and 12-week follow-ups. RESULTS: No significant changes in the SDS total score were observed for either group at each postintervention point. However, the IPC group showed a large effect size at the 4- and 12-week follow-ups. A significant intergroup difference was observed after 4 weeks. No significant intergroup difference was observed after 12 weeks, but there was a large effect size. CONCLUSIONS: IPC appeared to have effects at 4 weeks postintervention. Because this was an exploratory study, further research is necessary.

Increased prefrontal cortex activity on near-infrared spectroscopy after interpersonal counseling in individuals with attention deficit hyperactivity disorder.

[Purpose] The number of patients with attention deficit hyperactivity disorder has been increasing. These patients show low activity in the prefrontal cortex, which can be improved by pharmacotherapy and neurofeedback training. This exploratory study aimed to examine whether the hemodynamic response in the prefrontal cortex during an inhibition response in patients with attention deficit hyperactivity disorder tendencies increased after interpersonal counseling. [Participants and Methods] Participants (n=5) received three interpersonal counseling sessions. Interpersonal counseling focuses on the patient's current problems and devises specific coping strategies, and it can be performed by healthcare personnel such as physiotherapists. Prefrontal cortex activity during a suppression reaction task was measured by using near-infrared spectroscopy at baseline and post-interpersonal counseling. The outcome was a difference in the oxyhemoglobin level from baseline to post-interpersonal counseling. [Results] The oxyhemoglobin level in the prefrontal cortex significantly increased post-interpersonal counseling. [Conclusion] These results suggested that interpersonal counseling could improve the hemodynamic response in the prefrontal cortex under inhibition in individuals with attention deficit hyperactivity disorder tendencies, suggesting that interpersonal counseling may be effective for treating attention deficit hyperactivity disorder symptoms.

Special Features of COVID-19 in the FMODB: Fragment Molecular Orbital Calculations and Interaction Energy Analysis of SARS-CoV-2-Related Proteins.

SARS-CoV-2 is the causative agent of coronavirus (known as COVID-19), the virus causing the current pandemic. There are ongoing research studies to develop effective therapeutics and vaccines against COVID-19 using various methods and many results have been published. The structure-based drug design of SARS-CoV-2-related proteins is promising, however, reliable information regarding the structural and intra- and intermolecular interactions is required. We have conducted studies based on the fragment molecular orbital (FMO) method for calculating the electronic structures of protein complexes and analyzing their quantitative molecular interactions. This enables us to extensively analyze the molecular interactions in residues or functional group units acting inside the protein complexes. Such precise interaction data are available in the FMO database (FMODB) (https://drugdesign.riken.jp/FMODB/). Since April 2020, we have performed several FMO calculations on the structures of SARS-CoV-2-related proteins registered in the Protein Data Bank. We have published the results of 681 structures, including three structural proteins and 11 nonstructural proteins, on the COVID-19 special page (as of June 8, 2021). In this paper, we describe the entire COVID-19 special page of the FMODB and discuss the calculation results for various proteins. These data not only aid the interpretation of experimentally determined structures but also the understanding of protein functions, which is useful for rational drug design for COVID-19.

Regulation of Collective Metastasis by Nanolumenal Signaling.

Collective metastasis is defined as the cohesive migration and metastasis of multicellular tumor cell clusters. Disrupting various cell adhesion genes markedly reduces cluster formation and colonization efficiency, yet the downstream signals transmitted by clustering remain largely unknown. Here, we use mouse and human breast cancer models to identify a collective signal generated by tumor cell clusters supporting metastatic colonization. We show that tumor cell clusters produce the growth factor epigen and concentrate it within nanolumina-intercellular compartments sealed by cell-cell junctions and lined with microvilli-like protrusions. Epigen knockdown profoundly reduces metastatic outgrowth and switches clusters from a proliferative to a collective migratory state. Tumor cell clusters from basal-like 2, but not mesenchymal-like, triple-negative breast cancer cell lines have increased epigen expression, sealed nanolumina, and impaired outgrowth upon nanolumenal junction disruption. We propose that nanolumenal signaling could offer a therapeutic target for aggressive metastatic breast cancers.

Pluripotent stem cell-derived skeletal muscle fibers preferentially express myosin heavy-chain isoforms associated with slow and oxidative muscles.

BACKGROUND: Skeletal muscle function is essential for health, and it depends on the proper activity of myofibers and their innervating motor neurons. Each adult muscle is composed of different types of myofibers with distinct contractile and metabolic characteristics. The proper balance of myofiber types is disrupted in most muscle degenerative disorders, representing another factor compromising muscle function. One promising therapeutic approach for the treatment of these diseases is cell replacement based on the targeted differentiation of pluripotent stem cells (PSCs) towards the myogenic lineage. We have previously shown that transient induction of Pax3 or Pax7 in PSCs allows for the generation of skeletal myogenic progenitors endowed with myogenic regenerative potential, but whether they contribute to different fiber types remains unknown. RESULTS: Here, we investigate the fiber type composition of mouse PSC-derived myofibers upon their transplantation into dystrophic and non-dystrophic mice. Our data reveal that PSC-derived myofibers express slow and oxidative myosin heavy-chain isoforms, along with developmental myosins, regardless of the recipient background. Furthermore, transplantation of the mononuclear cell fraction re-isolated from primary grafts into secondary recipients results in myofibers that maintain preferential expression of slow and oxidative myosin heavy-chain isoforms but no longer express developmental myosins, thus indicating postnatal composition. CONCLUSIONS: Considering oxidative fibers are commonly spared in the context of dystrophic pathogenesis, this feature of PSC-derived myofibers could be advantageous for therapeutic applications.

Transplantation studies reveal internuclear transfer of toxic RNA in engrafted muscles of myotonic dystrophy 1 mice.

BACKGROUND: Stem cell transplantation represents a potential therapeutic option for muscular dystrophies (MD). However, to date, most reports have utilized mouse models for recessive types of MD. Here we performed studies to determine whether myotonic dystrophy 1 (DM1), an autosomal dominant type of MD, could benefit from cell transplantation. METHODS: We injected human pluripotent stem (PS) cell-derived myogenic progenitors into the muscles of a novel mouse model combining immunodeficiency and skeletal muscle pathology of DM1 and investigated transplanted mice for engraftment as well as for the presence of RNA foci and alternative splicing pattern. FINDINGS: Engraftment was clearly observed in recipient mice, but unexpectedly, we detected RNA foci in donor-derived engrafted myonuclei. These foci proved to be pathogenic as we observed MBNL1 sequestration and abnormal alternative splicing in donor-derived transcripts. INTERPRETATION: It has been assumed that toxic CUG repeat-containing RNA forms foci in situ in the nucleus in which it is expressed, but these data suggest that CUG repeat-containing RNA may also exit the nucleus and traffic to other nuclei in the syncytial myofiber, where it can exert pathological effects. FUND: This project was supported by funds from the LaBonte/Shawn family and NIH grants R01 AR055299 and AR071439 (R.C.R.P.). R.M-G. was funded by CONACyT-Mexico (#394378).

Pluripotent stem cell-derived myogenic progenitors remodel their molecular signature upon in vivo engraftment.

Optimal cell-based therapies for the treatment of muscle degenerative disorders should not only regenerate fibers but provide a quiescent satellite cell pool ensuring long-term maintenance and regeneration. Conditional expression of Pax3/Pax7 in differentiating pluripotent stem cells (PSCs) allows the generation of myogenic progenitors endowed with enhanced regenerative capacity. To identify the molecular determinants underlying their regenerative potential, we performed transcriptome analyses of these cells along with primary myogenic cells from several developmental stages. Here we show that in vitro-generated PSC-derived myogenic progenitors possess a molecular signature similar to embryonic/fetal myoblasts. However, compared with fetal myoblasts, following transplantation they show superior myofiber engraftment and ability to seed the satellite cell niche, respond to multiple reinjuries, and contribute to long-term regeneration. Upon engraftment, the transcriptome of reisolated Pax3/Pax7-induced PSC-derived myogenic progenitors changes toward a postnatal molecular signature, particularly in genes involved in extracellular matrix remodeling. These findings demonstrate that Pax3/Pax7-induced myogenic progenitors remodel their molecular signature and functionally mature upon in vivo exposure to the adult muscle environment.

The Effect of Interpersonal Counseling for Subthreshold Depression in Undergraduates: An Exploratory Randomized Controlled Trial.

BACKGROUND: Subthreshold depression and poor stress coping strategies are major public health problems among undergraduates. Interpersonal counseling (IPC) is a brief structured psychological intervention originally designed for use in primary care to treat depressive patients whose symptoms arose from current life stress. OBJECTIVES: This study examined the efficacy of IPC in treating subthreshold depression and coping strategies among undergraduates in school counseling. MATERIALS AND METHODS: We carried out an exploratory randomized controlled trial comparing the efficacy of IPC with counseling as usual (CAU). Participants were 31 undergraduates exhibiting depression without a psychiatric diagnosis. RESULTS: The Zung Self-Rating Depression Scale total score decreased significantly in the IPC group (n = 15; Z = -2.675, p = .007), but not in the CAU group (n = 16). The task-oriented coping score of the Coping Inventory for Stressful Situations showed a tendency towards a greater increase in the IPC group than in the CAU group (t = 1.919, df = 29, p = .065). CONCLUSIONS: The IPC might be more useful for student counseling because it can teach realistic coping methods and reduce depressive symptoms in a short period. Further studies using more participants are required.

Use of Interpersonal Counseling for Modern Type Depression.

A novel form of depression, called "modern type depression" (MTD), has been increasing in prevalence in Japan. Patients with MTD present with an overt appeal of depressive mood and a desire to be excused from their work duties; as such, this can cause considerable trouble in the workplace. Psychosocial interventions should be primarily considered for the treatment of MTD. Interpersonal counseling (IPC), which has proven effective for treating subthreshold depression, may be effective for MTD. However, IPC is rarely done in Japan. Herein, we report on a successful case of IPC for a woman in her thirties who was about to quit her job due to MTD (diagnosed by the criteria for research use). After IPC, the patient enjoyed good communication with her boss and continued her job without succumbing to her depression. This case suggests that IPC may be effective for MTD in workers and further highlights the benefits of teaching interpersonal communication methods in the workplace.

Generation of skeletal myogenic progenitors from human pluripotent stem cells using non-viral delivery of minicircle DNA.

Currently, the most efficient and promising approach for generating large numbers of engraftable human skeletal myogenic progenitors from pluripotent stem cells requires the conditional in vitro overexpression of PAX7 using lentiviral vectors. Because a non-integrating approach would be preferable to eliminate or minimize the risk associated with random genomic integration, here we investigate whether transient expression of PAX7 using minicircle DNA would enable the generation of functional pluripotent stem cell-derived myogenic progenitors, equivalent to those generated by lentivirus. Our results demonstrate that upon multiple transfections, the minicircle approach allows for the scalable generation of myogenic progenitors and these undergo efficient terminal differentiation in vitro. However, transplantation of minicircle-generated myogenic progenitors resulted in limited engraftment. This is probably due to less efficient delivery and more transient PAX7 expression in these cultures since PAX7 downregulation is accompanied by high level of spontaneous differentiation. Thus, although the in vitro data shows that the minicircle approach could potentially replace the use of lentivirus, improvements in the transfection/expression system will be necessary before it will be a feasible strategy for the generation of myogenic progenitors for cell replacement therapy.

PAX7 Targets, CD54, Integrin α9ß1, and SDC2, Allow Isolation of Human ESC/iPSC-Derived Myogenic Progenitors.

Pluripotent stem (PS)-cell-derived cell types hold promise for treating degenerative diseases. However, PS cell differentiation is intrinsically heterogeneous; therefore, clinical translation requires the development of practical methods for isolating progenitors from unwanted and potentially teratogenic cells. Muscle-regenerating progenitors can be derived through transient PAX7 expression. To better understand the biology, and to discover potential markers for these cells, here we investigate PAX7 genomic targets and transcriptional changes in human cells undergoing PAX7-mediated myogenic commitment. We identify CD54, integrin α9ß1, and Syndecan2 (SDC2) as surface markers on PAX7-induced myogenic progenitors. We show that these markers allow for the isolation of myogenic progenitors using both fluorescent- and CGMP-compatible magnetic-based sorting technologies and that CD54+α9ß1+SDC2+ cells contribute to long-term muscle regeneration in vivo. These findings represent a critical step toward enabling the translation of PS-cell-based therapies for muscle diseases.

Endoglin: a novel target for therapeutic intervention in acute leukemias revealed in xenograft mouse models.

Endoglin (CD105), a receptor of the transforming growth factor-ß superfamily, has been reported to identify functional long-term repopulating hematopoietic stem cells, and has been detected in certain subtypes of acute leukemias. Whether this receptor plays a functional role in leukemogenesis remains unknown. We identified endoglin expression on the majority of blasts from patients with acute myeloid leukemia (AML) and acute B-lymphoblastic leukemia (B-ALL). Using a xenograft model, we find that CD105+ blasts are endowed with superior leukemogenic activity compared with the CD105- population. We test the effect of targeting this receptor using the monoclonal antibody TRC105, and find that in AML, TRC105 prevented the engraftment of primary AML blasts and inhibited leukemia progression following disease establishment, but in B-ALL, TRC105 alone was ineffective due to the shedding of soluble CD105. However, in both B-ALL and AML, TRC105 synergized with reduced intensity myeloablation to inhibit leukemogenesis, indicating that TRC105 may represent a novel therapeutic option for B-ALL and AML.

Mechanism of microtubule lumen entry for the α-tubulin acetyltransferase enzyme αTAT1.

Microtubules are structural polymers inside of cells that are subject to posttranslational modifications. These posttranslational modifications create functionally distinct subsets of microtubule networks in the cell, and acetylation is the only modification that takes place in the hollow lumen of the microtubule. Although it is known that the α-tubulin acetyltransferase (αTAT1) is the primary enzyme responsible for microtubule acetylation, the mechanism for how αTAT1 enters the microtubule lumen to access its acetylation sites is not well understood. By performing biochemical assays, fluorescence and electron microscopy experiments, and computational simulations, we found that αTAT1 enters the microtubule lumen through the microtubule ends, and through bends or breaks in the lattice. Thus, microtubule structure is an important determinant in the acetylation process. In addition, once αTAT1 enters the microtubule lumen, the mobility of αTAT1 within the lumen is controlled by the affinity of αTAT1 for its acetylation sites, due to the rapid rebinding of αTAT1 onto highly concentrated α-tubulin acetylation sites. These results have important implications for how acetylation could gradually accumulate on stable subsets of microtubules inside of the cell.

Effectiveness of Simple Individual Psychoeducation for Bipolar II Disorder.

Several studies have proven the effectiveness of psychoeducation in bipolar II disorder patients; however, simpler psychoeducation is needed in daily medical practice. Therefore, we devised a simple individual psychoeducation program, which involved 20-minute sessions spent reading a textbook aloud in the waiting time before examination. Here, we report a successful case of simple individual psychoeducation with a patient with bipolar II disorder, a 64-year-old woman who had misconceptions surrounding her mood due to 24 years of treatment for depression. Her perception of mood state, particularly mixed state, was dramatically changed, and her quality of life was improved after the simple individual psychoeducation. This case suggests that the simple individual psychoeducation could be effective for bipolar II disorder by improving understanding of the disease and by meeting different individual needs.

Targeted mutagenesis of intergenic regions in the Neisseria gonorrhoeae gonococcal genetic island reveals multiple regulatory mechanisms controlling type IV secretion.

Gonococci secrete chromosomal DNA into the extracellular environment using a type IV secretion system (T4SS). The secreted DNA acts in natural transformation and initiates biofilm development. Although the DNA and its effects are detectable, structural components of the T4SS are present at very low levels, suggestive of uncharacterized regulatory control. We sought to better characterize the expression and regulation of T4SS genes and found that the four operons containing T4SS genes are transcribed at very different levels. Increasing transcription of two of the operons through targeted promoter mutagenesis did not increase DNA secretion. The stability and steady-state levels of two T4SS structural proteins were affected by a homolog of tail-specific protease. An RNA switch was also identified that regulates translation of a third T4SS operon. The switch mechanism relies on two putative stem-loop structures contained within the 5' untranslated region of the transcript, one of which occludes the ribosome binding site and start codon. Mutational analysis of these stem loops supports a model in which induction of an alternative structure relieves repression. Taken together, these results identify multiple layers of regulation, including transcriptional, translational and post-translational mechanisms controlling T4SS gene expression and DNA secretion.