HOXC6-Mediated miR-188-5p Expression Induces Cell Migration through the Inhibition of the Tumor Suppressor FOXN2.

Homeobox C6 (HOXC6) is a transcription factor that plays a role in the malignant progression of various cancers. However, the roles of HOXC6 and its regulatory mechanism remain unclear. In this study, we used microRNA (miRNA) regulatory networks to identify key regulatory interactions responsible for HOXC6-mediated cancer progression. In microarray profiling of miRNAs, the levels of miRNAs such as hsa-miR-188-5p, hsa-miR-8063, and hsa-miR-8064 were significantly increased in HOXC6-overexpressing cells. Higher positive expression rates of HOXC6 and miR-188-5p were observed in malignant cancer. We also found that HOXC6 significantly upregulated miR-188-5p expression. The underlying function of HOXC6-mediated miR-188-5p expression was predicted through TargetScan and the MiRNA Database. Overexpression of mir-188-5p inhibited the expression of forkhead box N2 (FOXN2), a tumor suppressor gene. Furthermore, in the luciferase assay, miR-188-5p bound to the 3'-UTR of FOXN2 and was mainly responsible for the dysregulation of FOXN2 expression. Silencing FOXN2 induced cell migration, and the effect of FOXN2 silencing was enhanced when the HOXC6/miR-188-5p axis was induced. These results suggest that HOXC6/miR-188-5p may induce malignant progression in cancer by inhibiting the activation of the FOXN2 signaling pathway.

Choline Acetyltransferase Induces the Functional Regeneration of the Salivary Gland in Aging SAMP1/Kl -/- Mice.

Salivary gland dysfunction induces salivary flow reduction and a dry mouth, and commonly involves oral dysfunction, tooth structure deterioration, and infection through reduced salivation. This study aimed to investigate the impact of aging on the salivary gland by a metabolomics approach in an extensive aging mouse model, SAMP1/Klotho -/- mice. We found that the salivary secretion of SAMP1/Klotho -/- mice was dramatically decreased compared with that of SAMP1/Klotho WT (+/+) mice. Metabolomics profiling analysis showed that the level of acetylcholine was significantly decreased in SAMP1/Klotho -/- mice, although the corresponding levels of acetylcholine precursors, acetyl-CoA and choline, increased. Interestingly, the mRNA and protein expression of choline acetyltransferase (ChAT), which is responsible for catalyzing acetylcholine synthesis, was significantly decreased in SAMP1/Klotho -/- mice. The overexpression of ChAT induced the expression of salivary gland functional markers (α-amylase, ZO-1, and Aqua5) in primary cultured salivary gland cells from SAMP1/Klotho +/+ and -/- mice. In an in vivo study, adeno-associated virus (AAV)-ChAT transduction significantly increased saliva secretion compared with the control in SAMP1/Klotho -/- mice. These results suggest that the dysfunction in acetylcholine biosynthesis induced by ChAT reduction may cause impaired salivary gland function.

Exopolysaccharide from Lactobacillus plantarum HY7714 Protects against Skin Aging through Skin-Gut Axis Communication.

Skin aging occurs inevitably as a natural result of physiological changes over time. In particular, solar exposure of the skin accounts for up to 90% of skin damage. Numerous studies have examined the ability of dietary constituents to prevent skin aging, and recent research has emphasized the role of functional probiotics in intestinal function and skin aging. However, the mechanism of the interactions between aging and probiotics has not been elucidated yet. The aim of this study was to determine the role of exopolysaccharides (EPS) produced by lactic acid bacteria (LAB) identified as Lactobacillus plantarum HY7714 in regulating tight junctions in intestinal epithelial cells and increasing moisture retention in human dermal fibroblasts cells. We observed that HY7714 EPS controlled intestinal tight junctions in Caco-2 cells by upregulating the genes encoding occludin-1 (OCL-1) and zonula occluden-1 (ZO-1). In addition, HY7714 EPS effectively improved UVB-induced cytotoxicity and hydration capacity in HS68 cells by downregulating production of metalloproteinases (MMPs) and reactive oxygen species (ROS). In summary, HY7714 EPS is an effective anti-aging molecule in skin and may have therapeutic potential against skin diseases and UVB-induced damage. Therefore, HY7714 EPS serves as a functional substance in skin-gut axis communication.

Global analysis of gene expression profiles in the submandibular salivary gland of klotho knockout mice.

Salivary dysfunction commonly occurs in many older adults and is considered a physiological phenomenon. However, the genetic changes in salivary glands during aging have not been characterized. The present study analyzed the gene expression profile in salivary glands from accelerated aging klotho deficient mice (klotho-/-, 4 weeks old). Microarray analysis showed that 195 genes were differentially expressed (z-score > 2 in two independent arrays) in klotho null mice compared to wild-type mice. Importantly, alpha2-Na+ /K+ -ATPase (Atp1a2), Ca2+ -ATPase (Atp2a1), epidermal growth factor (EGF), and nerve growth factor (NGF), which have been suggested to be regulators of submandibular salivary gland function, were significantly decreased. When a network was constructed from the differentially expressed genes, proliferator-activated receptor-γ (PPAR γ), which regulates energy homeostasis and insulin sensitivity, was located at the core of the network. In addition, the expression of genes proposed to regulate various PPAR γ-related cellular pathways, such as Klk1b26, Egfbp2, Cox8b, Gpx3, Fabp3, EGF, and NGFß, was altered in the submandibular salivary glands of klotho-/- mice. Our results may provide clues for the identification of novel genes involved in salivary gland dysfunction. Further characterization of these differentially expressed genes will be useful in elucidating the genetic basis of aging-related changes in the submandibular salivary gland.

New metformin derivative HL156A prevents oral cancer progression by inhibiting the insulin-like growth factor/AKT/mammalian target of rapamycin pathways.

Metformin is a biguanide widely prescribed as an antidiabetic drug for type 2 diabetes mellitus patients. The purpose of the present study was to observe the effects of the new metformin derivative, HL156A, on human oral cancer cell and to investigate its possible mechanisms. It was observed that HL156A significantly decreased FaDu and YD-10B cell viability and colony formation in a dose-dependent way. HL156A also markedly reduced wound closure and migration of FaDu and YD-10B cells. We observed that HL156A decreased mitochondrial membrane potential and induced reactive oxygen species (ROS) levels and apoptotic cells with caspase-3 and -9 activation. HL156A inhibited the expression and activation of insulin-like growth factor (IGF)-1 and its downstream proteins, AKT, mammalian target of rapamycin (mTOR), and ERK1/2. In addition, HL156A activated AMP-activated protein kinase/nuclear factor kappa B (AMPK-NF-κB) signaling of FaDu and YD-10B cells. A xenograft mouse model further showed that HL156A suppressed AT84 mouse oral tumor growth, accompanied by down-regulated p-IGF-1, p-mTOR, proliferating cell nuclear antigen (PCNA) and promoted p-AMPK and TUNEL expression. These results suggest the potential value of the new metformin derivative HL156A as a candidate for a therapeutic modality for the treatment of oral cancer.

Hyaluronan suppresses lidocaine-induced apoptosis of human chondrocytes in vitro by inhibiting the p53-dependent mitochondrial apoptotic pathway.

AIM: Intra-articular injection of local anesthetics (LAs) is a common procedure for therapeutic purposes. However, LAs have been found toxic to articular cartilage, and hyaluronan may attenuate this toxicity. In this study we investigated whether hyaluronan attenuated lidocaine-induced chondrotoxicity, and if so, to elucidate the underlying mechanisms. METHODS: Human chondrocyte cell line SW1353 and newly isolated murine chondrocytes were incubated in culture medium containing hyaluronan and/or lidocaine for 72 h. Cell viability was evaluated using MTT assay. Cell apoptosis was detected with DAPI staining, caspase 3/7 activity assay and flow cytometry. Cell cycle distributions, ROS levels and mitochondrial membrane potential (ΔΨm) were determined using flow cytometry. The expression of p53 and p53-regulated gene products was measured with Western blotting. RESULTS: Lidocaine (0.005%-0.03%) dose-dependently decreased the viability of SW1353 cells. This local anesthetic (0.015%, 0.025%) induced apoptosis, G2/M phase arrest and loss of ΔΨm, and markedly increased ROS production in SW1353 cells. Hyaluronan (50-800 µg/mL) alone did not affect the cell viability, but co-treatment with hyaluronan (200 µg/mL) significantly attenuated lidocaine-induced apoptosis and other abnormalities in SW1353 cells. Furthermore, co-treatment with lidocaine and hyaluronan significantly decreased the levels of p53 and its transcription targets Bax and p21 in SW1353 cells, although treatment with lidocaine alone did not significantly change these proteins. Similar results were obtained in ex vivo cultured murine chondrocytes. CONCLUSION: Hyaluronan suppresses lidocaine-induced apoptosis of human chondrocytes in vitro through inhibiting the p53-dependent mitochondrial apoptotic pathway.

Crystal structure of the Golgi casein kinase.

The family with sequence similarity 20 (Fam20) kinases phosphorylate extracellular substrates and play important roles in biomineralization. Fam20C is the Golgi casein kinase that phosphorylates secretory pathway proteins within Ser-x-Glu/pSer motifs. Mutations in Fam20C cause Raine syndrome, an osteosclerotic bone dysplasia. Here we report the crystal structure of the Fam20C ortholog from Caenorhabditis elegans. The nucleotide-free and Mn/ADP-bound structures unveil an atypical protein kinase-like fold and highlight residues critical for activity. The position of the regulatory αC helix and the lack of an activation loop indicate an architecture primed for efficient catalysis. Furthermore, several distinct elements, including the presence of disulfide bonds, suggest that the Fam20 family diverged early in the evolution of the protein kinase superfamily. Our results reinforce the structural diversity of protein kinases and have important implications for patients with disorders of biomineralization.

BAG3 affects the nucleocytoplasmic shuttling of HSF1 upon heat stress.

Bcl2-associated athoanogene (BAG) 3 is a member of the co-chaperone BAG family. It is induced by stressful stimuli such as heat shock and heavy metals, and it regulates cellular adaptive responses against stressful conditions. In this study, we identified a novel role for BAG3 in regulating the nuclear shuttling of HSF1 during heat stress. The expression level of BAG3 was induced by heat stress in HeLa cells. Interestingly, BAG3 rapidly translocalized to the nucleus upon heat stress. Immunoprecipitation assay showed that BAG3 interacts with HSF1 under normal and stressed conditions and co-translocalizes to the nucleus upon heat stress. We also demonstrated that BAG3 interacts with HSF1 via its BAG domain. Over-expression of BAG3 down-regulates the level of nuclear HSF1 by exporting it to the cytoplasm during the recovery period. Depletion of BAG3 using siRNA results in reduced nuclear HSF1 and decreased Hsp70 promoter activity. BAG3 in MEF(hsf1(-/-)) cells actively translocalizes to the nucleus upon heat stress suggesting that BAG3 plays a key role in the processing of the nucleocytoplasmic shuttling of HSF1 upon heat stress.

Soluble klotho induces the heat shock factor 1 through EGR1 expression.

Klotho is an antiaging protein that has multiple functions. The purpose of this study is to investigate whether soluble klotho plays a role in cellular stress response pathways. We found that klotho deficiency (kl-/-) largely decreased HSF1 levels and impaired heat shock protein expression. Interestingly, recombinant soluble klotho-induced HSF1 and HSPs such as HSP90, HSP70, and HSP27 in kl-/- mouse embryonic fibroblasts (MEFs). Soluble Klotho treatment also induced cell proliferation and HSF1 promoter activity in MEF kl-/- cells in a concentration-dependent manner. Furthermore, using point mutagenesis, we identified regulatory/binding sites of transcription factors EGR1 regulated by soluble klotho in the HSF1 promoter. Taken together, our findings unravel the molecular basis of klotho and provide molecular evidence supporting a direct interaction between soluble klotho and HSF1-mediated stress response pathway.

Long-Term Enhancement of Botulinum Toxin Injections for Post-Stroke Spasticity by Use of Stretching Exercises-A Randomized Controlled Trial.

Botulinum toxin A (BONT/A) injections play a central role in the treatment of upper limb spasticity in stroke patients. We proposed structured stretching exercises to enhance the effect of post-stroke spasticity relief of the upper limbs following BONT/A injections. A total of 43 patients who had a stroke with grade 2 spasticity or higher on the Modified Ashworth Scale (MAS) in their upper-limb muscles were randomly assigned to the intervention (n = 21) or control group (n = 22). The former received structured stretching exercises after their BONT/A injections for 20 min, 5 days per week, for 6 months at a hospital, while the others conducted self-stretching exercises at home. The outcome measures were assessed before the intervention (T0) and after three (T1) and six months (T2). Significantly greater improvements in the MAS scores of the elbows, wrists, and fingers were found in the intervention group's patients at T1 and T2. The behavioral outcome measures, including shoulder pain, activities of daily living, and quality of life, and our electrophysiological studies also showed a significantly higher enhancement in this patient group. In conclusion, the structured stretching exercises plus BONT/A injections for six months showed a superior effect in relieving post-stroke upper-limb spasticity compared to self-stretching exercises.

Toe Grip Strength Is Associated with Improving Gait Function in Patients with Subacute Stroke.

Toe grip strength has recently been suggested to play an essential role in maintaining balance and postural stability for ambulatory function in older populations. This study aimed to investigate its association with improving gait function three months after onset in patients with subacute stroke. This longitudinal cohort study included 98 first-ever stroke patients (67 ± 9 years, 56% female) within one month from the onset who could not ambulate independently. Functional outcome indicators, including toe grip strength, hand grip strength, knee extensor strength, Fugl-Meyer Assessment of Lower Extremity (FMA_LE), and the Postural Assessment Scale for Stroke (PASS), were assessed before and three months after the intervention. We analyzed the correlation between participants' gait function using a 10-meter walk test time and various functional indicators. Then, multiple linear regression analysis was used to investigate whether toe grip strength was related to the improvement of gait function. Correlation analysis revealed a significant positive correlation between the 10MWT time and toe grip strength ratio (affected/unaffected side), with a moderate effect size (r = -0.61, p <0.001). Multiple regression analysis with covariates showed a significant relationship between 10MWT time and toe grip strength ratio (ß = -0.113, p < 0.001), FMA_LE (ß = -1.315, p = 0.004), PASS (ß = -3.275, p <0.001), and age (ß = -0.159, p = 0.004). In conclusion, toe grip strength was an essential factor associated with ambulatory function improvement in subacute stroke patients three months after onset. Additional toe grip muscle strengthening rehabilitation treatment can be expected to help improve the ambulatory function of subacute stroke patients in the future.

Modulus-tunable multifunctional hydrogel ink with nanofillers for 3D-Printed soft electronics.

Seamless integration and conformal contact of soft electronics with tissue surfaces have emerged as major challenges in realizing accurate monitoring of biological signals. However, the mechanical mismatch between the electronics and biological tissues impedes the conformal interfacing between them. Attempts have been made to utilize soft hydrogels as the bioelectronic materials to realize tissue-comfortable bioelectronics. However, hydrogels have several limitations in terms of their electrical and mechanical properties. In this study, we present the development of a 3D-printable modulus-tunable hydrogel with multiple functionalities. The hydrogel has a cross-linked double network, which greatly improves its mechanical properties. Functional fillers such as XLG or functionalized carbon nanotubes (fCNT) can be incorporated into the hydrogel to provide tunable mechanics (Young's modulus of 10-300 kPa) and electrical conductivity (electrical conductivity of ∼20 S/m). The developed hydrogel exhibits stretchability (∼1000% strain), self-healing ability (within 5 min), toughness (400-731 kJ/m3) viscoelasticity, tissue conformability, and biocompatibility. Upon examining the rheological properties in the modulated region, hydrogels can be 3D printed to customize the shape and design of the bioelectronics. These hydrogels can be fabricated into ring-shaped strain sensors for wearable sensor applications.

HOXC6 is deregulated in human head and neck squamous cell carcinoma and modulates Bcl-2 expression.

Homeobox C6 (HOXC6) genes belong to the homeoprotein family of transcription factors, which play an important role in morphogenesis and cellular differentiation during embryonic development. The aim of this study was to explore the role of HOXC6 in the regulation of Bcl-2 in human head and neck squamous cell carcinoma (HNSCC). The HOXC6 and Bcl-2 gene were identified as being overexpressed in HNSCC tissue and cell lines. Transfection assays demonstrated that HOXC6 increased the levels of Bcl-2 mRNA and protein. A luciferase reporter assay suggested that HOXC6 induced activity of the Bcl-2 promoter. A series of Bcl-2 promoter deletion mutants were examined and the minimal HOXC6-responsive region was identified to be in the TAAT motif (-420 bp) of the Bcl-2 promoter. Interestingly, the inhibition of HOXC6 using siRNA led to the repression of Bcl-2 expression and induced caspase-3-dependent apoptosis; overexpression of HOXC6 in HNSCC cells increased the resistance to paclitaxel-induced apoptosis. Together, our findings suggest that HOXC6 is an important mechanism of the anti-apoptotic pathway via regulation of Bcl-2 expression.

Radiologic changes of long term foot insole use in symptomatic pediatric flatfoot.

Clinically, flatfoot, known as pes planus, is quite prevalent. It is classified into 2 types: flexible and rigid, both of which may or may not have symptoms. If a flexible flatfoot is symptomatic, it must be treated to prevent subsequent complications. In principle, most physicians initially use conservative methods, such as foot insoles. This study aimed to demonstrate the effect of long term use of a foot insole using plain radiography as an objective measurement in children with symptomatic flexible flatfoot (SFFF) in large samples. This study analyzed the medical records of 292 children aged < 18 years who were diagnosed with SFFF. Of these, 200 children (62 boys and 138 girls, mean age: 6.49 ± 2.96 years) were selected and conservatively treated with foot insoles. They were periodically followed up within 3 to 4 months to modify the foot insole and perform radiologic evaluations, such as foot radiography. The calcaneal pitch angle (CPA) and talo first metatarsal angle were measured and compared individually using foot lateral radiographs, which were pictured in a bilateral barefoot state. The treatment was terminated by repeating the same procedure until the symptoms disappeared. A significant improvement (P < .001) was observed in the radiological indicators, both CPA and talo first metatarsal angle, regardless of age, after the application of soft foot insoles. However, the right foot CPA in the group with valgus deformity was an exception (P = .078). This study showed that in children diagnosed with SFFF under 18 years of age, wearing a periodically revised foot insole as conservative treatment could not only decrease the symptoms, but also improve the radiologic indices.

3D printing of mechanically tough and self-healing hydrogels with carbon nanotube fillers.

Hydrogels have the potential to play a crucial role in bioelectronics, as they share many properties with human tissues. However, to effectively bridge the gap between electronics and biological systems, hydrogels must possess multiple functionalities, including toughness, stretchability, self-healing ability, three-dimensional (3D) printability, and electrical conductivity. Fabricating such tough and self-healing materials has been reported, but it still remains a challenge to fulfill all of those features, and in particular, 3D printing of hydrogel is in the early stage of the research. In this paper, we present a 3D printable, tough, and self-healing multi-functional hydrogel in one platform made from a blend of poly(vinyl alcohol) (PVA), tannic acid (TA), and poly(acrylic acid) (PAA) hydrogel ink (PVA/TA/PAA hydrogel ink). Based on a reversible hydrogen-bond (H-bond)-based double network, the developed 3D printable hydrogel ink showed excellent printability via shear-thinning behavior, allowing high printing resolution (~100 µm) and successful fabrication of 3D-printed structure by layer-by-layer printing. Moreover, the PVA/TA/PAA hydrogel ink exhibited high toughness (tensile loading of up to ~45.6 kPa), stretchability (elongation of approximately 650%), tissue-like Young's modulus (~15 kPa), and self-healing ability within 5 min. Furthermore, carbon nanotube (CNT) fillers were successfully added to enhance the electrical conductivity of the hydrogel. We confirmed the practicality of the hydrogel inks for bioelectronics by demonstrating biocompatibility, tissue adhesiveness, and strain sensing ability through PVA/TA/PAA/CNT hydrogel ink.

Three-Dimensional Magnetic Rehabilitation, Robot-Enhanced Hand-Motor Recovery after Subacute Stroke: A Randomized Controlled Trial.

We developed an end-effector-type rehabilitation robot that can uses electro- and permanent magnets to generate a three-way magnetic field to assist hand movements and perform rehabilitation therapy. This study aimed to investigate the therapeutic effect of a rehabilitation program using a three-dimensional (3D) magnetic force-based hand rehabilitation robot on the motor function recovery of the paralyzed hands of patients with stroke. This was a double-blind randomized controlled trial in which 36 patients with subacute stroke were assigned to intervention and control groups of 18 patients each. The intervention group received 30 min of rehabilitation therapy per day for a month using a 3D magnetic force-driven hand rehabilitation robot, whereas the control group received 30 min of conventional occupational therapy to restore upper-limb function. The patients underwent three behavioral assessments at three time points: before starting treatment (T0), after 1 month of treatment (T1), and at the follow-up 1-month after treatment completion (T2). The primary outcome measure was the Wolf Motor Function Test (WMFT), and secondary outcome measures included the Fugl-Meyer Assessment of the Upper Limb (FMA_U), Modified Barthel Index (MBI), and European Quality of Life Five Dimensions (EQ-5D) questionnaire. No participant safety issues were reported during the intervention. Analysis using repeated measures analysis of variance showed significant interaction effects between time and group for both the WMFT score (p = 0.012) and time (p = 0.010). In post hoc analysis, the WMFT scores and time improved significantly more in the patients who received robotic rehabilitation at T1 than in the controls (p = 0.018 and p = 0.012). At T2, we also consistently found improvements in both the WMFT scores and times for the intervention group that were superior to those in the control group (p = 0.024 and p = 0.018, respectively). Similar results were observed for FMA_U, MBI, and EQ-5D. Rehabilitation using the 3D hand-rehabilitation robot effectively restored hand function in the patients with subacute stroke, contributing to improvement in daily independence and quality of life.

A Mechanically Resilient and Tissue-Conformable Hydrogel with Hemostatic and Antibacterial Capabilities for Wound Care.

Hydrogels are used in wound dressings because of their tissue-like softness and biocompatibility. However, the clinical translation of hydrogels remains challenging because of their long-term stability, water swellability, and poor tissue adhesiveness. Here, tannic acid (TA) is introduced into a double network (DN) hydrogel consisting of poly(vinyl alcohol) (PVA) and poly(acrylic acid) (PAA) to realize a tough, self-healable, nonswellable, conformally tissue-adhesive, hemostatic, and antibacterial hydrogel. The TA within the DN hydrogel forms a dynamic network, enabling rapid self-healing (within 5 min) and offering effective energy dissipation for toughness and viscoelasticity. Furthermore, the hydrophobic moieties of TA provide a water-shielding effect, rendering the hydrogel nonswellable. A simple chemical modification to the hydrogel further strengthens its interfacial adhesion with tissues (shear strength of ≈31 kPa). Interestingly, the TA also can serve as an effective hemostatic (blood-clotting index of 58.40 ± 1.5) and antibacterial component, which are required for a successful wound dressing. The antibacterial effects of the hydrogel are tested against Escherichia coli and Staphylococcus aureus. Finally, the hydrogel is prepared in patch form and applied to a mouse model to test in vivo biocompatibility and hemostatic performances.

Intrinsically Nonswellable Multifunctional Hydrogel with Dynamic Nanoconfinement Networks for Robust Tissue-Adaptable Bioelectronics.

Developing bioelectronics that retains their long-term functionalities in the human body during daily activities is a current critical issue. To accomplish this, robust tissue adaptability and biointerfacing of bioelectronics should be achieved. Hydrogels have emerged as promising materials for bioelectronics that can softly adapt to and interface with tissues. However, hydrogels lack toughness, requisite electrical properties, and fabrication methodologies. Additionally, the water-swellable property of hydrogels weakens their mechanical properties. In this work, an intrinsically nonswellable multifunctional hydrogel exhibiting tissue-like moduli ranging from 10 to 100 kPa, toughness (400-873 J m-3 ), stretchability (≈1000% strain), and rapid self-healing ability (within 5 min), is developed. The incorporation of carboxyl- and hydroxyl-functionalized carbon nanotubes (fCNTs) ensures high conductivity of the hydrogel (≈40 S m-1 ), which can be maintained and recovered even after stretching or rupture. After a simple chemical modification, the hydrogel shows tissue-adhesive properties (≈50 kPa) against the target tissues. Moreover, the hydrogel can be 3D printed with a high resolution (≈100 µm) through heat treatment owing to its shear-thinning capacity, endowing it with fabrication versatility. The hydrogel is successfully applied to underwater electromyography (EMG) detection and ex vivo bladder expansion monitoring, demonstrating its potential for practical bioelectronics.

Heat shock factor 4a (HSF4a) represses HSF2 expression and HSF2-mediated transcriptional activity.

Heat shock factors (HSFs) are the main transcriptional regulators of the stress-induced expression of heat shock protein genes. HSF2, which is one of the HSFs, is activated during differentiation and development but it is unclear how they regulate during cellular processes. Here, we examined the role of HSF4a on the regulation of HSF2 in HEK 293 cells. We found that HSF2 levels are negatively correlated with HSF4a expression and that overexpression of HSF4a reduces hemin-induced HSF2 mRNA and protein levels. Moreover, hemin-induced activation of HSF2 was also markedly inhibited in HSF4a expressed cells. Immunoprecipitation assay showed that HSF2 binds to the oligomerization domain of HSF4a. Hemin treatment inhibited their interaction and induced localization of HSF2 and HSF4a in nuclear. In addition, we found that HSF4a or HSF4a DNA binding domain (117 aa) inhibited the activity of hemin-induced HSP70 promoter. Consequently, HSF4a inhibits HSF2 expression or transcriptional activity through negative regulation of HSF2 binding to the HSP70 promoter. In summary, our findings suggest novel mechanisms of HSF2 regulation controlled by HSF4a.

Homeobox C5 expression is associated with the progression of 4-nitroquinoline 1-oxide-induced rat tongue carcinogenesis.

BACKGROUND: Aberrant expression of homeobox genes (HOX), normally required for the differentiation of a particular tissue, has been reported in several types of cancer, but poorly addressed in oral squamous cell carcinoma (OSCC). The present study investigated the expression of HOXC5 in OSCC and identified molecular biomarker whose expression is associated with the multistep oral carcinogenesis. METHODS: The expression of HOXC5, proliferation cell nuclear antigen (PCNA), and Bcl-2 was examined by RT-PCR and Western blot analysis and confirmed by immunohistochemistry and transferase-mediated dUTP nick end-labeling (TUNEL) assay in a 4-nitroquinoline 1-oxide (4NQO)-induced rat tongue carcinogenesis model. RESULTS: Homeobox genes C5 was overexpressed in SCC tissues, but not in normal tissues by RT-PCR and Western blot analysis. Along with the progress of multistep carcinogenesis, the levels of HOXC5 expression of mRNA and protein significantly increased during the dysplasia (moderate to severe dysplasia) when compared with normal and hyperplasia. The levels of PCNA and Bcl-2 were sequentially increased from hyperplasia to dysplasia and SCC. By immunohistochemistry, HOXC5 expression was significantly increased in dysplasia, whereas PCNA expression was gradually increased during tongue carcinogenesis. TUNEL-positive cells were increased until dysplasia, but reduced in SCC. CONCLUSIONS: These results indicate that overexpression of HOXC5 is correlated with oral carcinogenesis and strongly contributed to the development of OSCC. HOXC5 may be a useful biomarker and has an emerging therapeutic target of OSCC.