Static reconstruction of malar region in facial paralysis: a new alternative technique for plasty of symmetric mouth appearance.

Static suspension using fascia lata graft is used as a reconstructive procedure against drooping of the mouth corner for treating longstanding facial paralysis. Although it achieves symmetry at rest, movement of the mouth corner at mouth opening is restricted to some extent because it is fixed with fascia lata to the immovable temporal fascia, the parotid fascia, or bones. This was overcome by suspending the mouth corner to the mandibular coronoid process with fascia lata, which enabled a shift of the mouth corner with mouth opening and closure. The nine patients discussed in this study were operated on since 1994 for longstanding facial paralysis and followed-up for over 1.5 years. As in conventional static suspension, the fascia lata was harvested and split into two bands. Next, one semi-oval fascial loop was inserted around the paralysed part of the mouth and tied with another fascial band at the mouth corner, which was looped to the mandibular coronoid process. The suspended fascia lata graft was relaxed with anteroinferior movement of the coronoid process at mouth opening, enabling the mouth corner to shift inferiorly. The mouth corner returned to its original position at mouth closure, and the nasolabial fold deepened during mastication. No limitation in mouth opening was observed. Suspension of the mouth corner to the mandibular coronoid process provided a dynamic element, thereby restoring a near-normal shift. The procedure is considered as an alternative for reconstructing the malar region of patients with facial paralysis and in whom dynamic reconstruction is not indicated.

Depletion of Dnmt1-associated protein 1 triggers DNA damage and compromises the proliferative capacity of hematopoietic stem cells.

Dnmt1-associated protein 1 (Dmap1) is a core component of the NuA4 histone acetyltransferase complex and the Swr1 chromatin-remodeling complex. However, the cellular function of Dmap1 remains largely unknown. We previously reported that Dmap1 plays a crucial role in DNA repair and is indispensable for the maintenance of chromosomal integrity of mouse embryonic fibroblasts. In this study, we examined the role of Dmap1 in self-renewing HSCs. Dmap1-knockdown induced by Dmap1-specific shRNA severely compromised the proliferative capacity of HSCs in vitro and long-term repopulating capacity of HSCs in recipient mice. Dmap1-knockdown in HSCs triggered DNA damage as evident by the formation of foci of gamma-H2AX and activated p53-dependent cell cycle checkpoints. Deletion of p53 in HSCs abrogated the activation of p53-dependent cell cycle checkpoints, but did not restore the HSC function impaired by the knockdown of Dmap1. These findings suggest that Dmap1 is essential for the maintenance of genomic integrity of self-renewing HSCs and highlight DNA damage as one of the major stresses causing HSC depletion.

Treatment of velopharyngeal inadequacy in a patient with submucous cleft palate and myasthenia gravis.

OBJECTIVE: To describe the clinical course and management of a patient with submucous cleft palate who developed myasthenia gravis (MG) as an adult and suffered recurrent hypernasality. Few reports have described MG patients undergoing pharyngeal flap surgery for velopharyngeal incompetence, and these have described only slight speech improvement in such patients. DESIGN: Case report. PATIENT: The patient underwent primary pushback palatoplasty and superiorly based pharyngeal flap surgery for submucous cleft and short palate at age 7. Hypernasality showed major improvement after initial surgery. At age 19, the patient developed MG that triggered the recurrence of velopharyngeal incompetence. INTERVENTION: After MG was treated, revision pushback palatoplasty was performed for velopharyngeal incompetence when the patient was 24 years old. Preoperatively and postoperatively, the patient was evaluated by the same speech-language-hearing therapists, each with at least 5 years of clinical experience in cleft palate speech. RESULTS: After the second pushback palatoplasty, hypernasality and audible nasal air emission during speech decreased to mild. CONCLUSION: Primary pushback palatoplasty and pharyngeal flap surgery were performed for the submucous cleft palate. Revision pushback palatoplasty improved velopharyngeal inadequacy induced by MG. Decreased perceived nasality positively influenced the patient's quality of life. Combined pushback palatoplasty and pharyngeal flap surgery is thus an option in surgical treatment for velopharyngeal inadequacy to close the cleft and the velopharyngeal orifice in cases of cleft palate and MG.

Cyclical cell stretching of skin-derived fibroblasts downregulates connective tissue growth factor (CTGF) production.

Delayed healing of skin wounds can be caused by wound instability, whereas appropriate massage or exercise prevents sclerosis and scar contracture. However, the mechanism by which wound healing is related to mechanical stress has not been fully elucidated. The present study aimed to identify whether mechanical stretching of fibroblasts reduces their production of extracellular matrix. We transferred skin fibroblasts into collagen-coated elastic silicone chambers, cultured them on a stretching apparatus, and used RT-PCR to examine the effects of mechanical stretching on the expression levels of 17 genes related to extracellular matrix production and growth factor secretion. We found that connective tissue growth factor (CTGF) was downregulated after 24 hr of cell stretching. Specifically, the CTGF mRNA and protein levels were 50% and 48% of the control levels, respectively. These findings suggest that cyclic stretching of fibroblasts contributes to anti-fibrotic processes by reducing CTGF production.

Expressions of androgen receptor and its co-regulators in carcinoma ex pleomorphic adenoma of salivary gland.

AIMS: Androgen receptor (AR) signalling is involved in cancer progression. The expression of AR has been reported in carcinoma ex pleomorphic adenoma (CXPA) of salivary gland, however AR gene status and the expressions of cofactors for AR signalling have not been investigated. The aims of this study were to investigate the expressions of each of the molecules that contribute to AR activation with or without ligands in CXPA. In addition, AR gene amplification and single-nucleotide polymorphism were investigated. METHODS: Ten cases of CXPA and 23 cases of pleomorphic adenomas (PA) of the salivary glands were immunostained for the AR co-regulators (SRC1, p300, and NCoR1) and the signalling molecules involved in the ligand-independent pathway (i.e., HER-2/neu and STAT3). AR gene amplification and single-nucleotide polymorphism were investigated by dual-coloured fluorescent in situ hybridisation and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), respectively. RESULTS: AR expression was observed in nine of 10 cases of CXPA and in 30.4% of PA cases, a statistically significant difference. The expression, with low or high intensity, of HER-2/neu and STAT3 was more frequent in CXPA (6/10 and 9/10, respectively) than in PA (0% and 46.7%). The expression of co-activators was also stronger, though only slightly, in CXPA than in PA. The gain of chromosome X and AR gene amplification were not observed in any CXPA or PA cases, and the G --> A allele in codon 211 was detected in only one case (a CXPA). CONCLUSIONS: These results suggest that although AR may be activated in the pathway with or without ligands, the expression of co-regulators and AR gene aberrations are not involved in the carcinogenesis of CXPA.

Hemi aplasia of the nose with complete cleft lip and palate of the contralateral side.

Nasal aplasia, including hemi aplasia of the nose, is a rare congenital anomaly of the nose. Since the ipsilateral side tends to be affected more frequently than the contralateral side of the face in half nose anomalies, only reports concerning the ipsilateral defect are numerous. This report presents an unusual case of hemi aplasia of the nose with complete cleft lip and palate of the contralateral side. A local flap on the nasal dorsum was used for nasal reconstruction, where correction of the elongation of the inner canthal distance and the shape of the inner canthus was performed.

DNA methylation changes during cleft palate formation induced by retinoic acid in mice.

OBJECTIVES: The aim of this study was to analyze epigenetic (specifically, DNA methylation) participation in the mechanisms of cleft palate only induced by maternal exposure to all-trans retinoic acid in mice. DESIGN: Cleft palate only was induced in fetuses by maternal exposure to all-trans retinoic acid. Their secondary palates were excised for analysis. Cytosine extension assay and restriction landmark genomic scanning were performed to analyze DNA methylation status. The expression levels of the DNA methyltransferases were examined by real-time reverse transcriptase-polymerase chain reaction. RESULTS: Using cytosine extension assay, on gestation day 14.5, the status of DNA methylation within CpG islands and in global DNA was decreased significantly in all-trans retinoic acid-treated groups compared with the controls (p < .01 and p < .05). In the controls, the status within CpG islands on gestation day 14.5 was significantly increased compared with gestation days 13.5 and 18.5 (p < .01). Using real-time reverse transcriptase-polymerase chain reaction, there was no significant change in the expression of DNA methyltransferases, except on gestation day 18.5. Using restriction landmark genomic scanning on gestation day 18.5, five spots (0.49%) in the controls and one spot (0.1%) in all-trans retinoic acid-treated groups were specifically detected. CONCLUSIONS: These results indicate that changes in DNA methylation may play an important role in the manifestation of cleft palate only caused by environmental factors such as maternal exposure to all-trans retinoic acid.

Tessier number 7 cleft with oblique clefts of bilateral soft palates and rare symmetric structure of zygomatic arch.

Craniofacial clefts are very rare congenital deformities. Tessier's numbering classification is a simple system universally accepted. Excluding cleft lip and palate, the transverse or lateral cleft (Tessier number 7) is the most common type of craniofacial cleft, manifesting macrostomia without skeletal abnormalities. We present a case of Tessier number 7 cleft with oblique clefting of the soft palate bilaterally and rare symmetric structures of the zygomatic arch.

Low-dose GH supplementation reduces the TLR2 and TNF-alpha expressions in visceral fat.

The increased population of TLR2/TNF-alpha co-expressing adipocytes is associated with the development of insulin resistance. We have herein shown the significance of low-dose growth hormone (GH) supplementation for the regulation of TLR2 and TNF-alpha expressions in visceral fat using different kinds of mouse models fed with a high-fat diet. Low-dose GH supplementation reduced the increased population of TLR2/TNF-alpha co-expressing adipocytes in high-fat fed mice. The neutralization of IGF-1 abolished the effect of GH supplementation on the TLR2 expression using GH-overexpressing mice. IGF-1, but not GH, inhibited the FFA-induced TLR2 and TNF-alpha expression in 3T3-L1 cells. Finally, low-dose GH supplementation reduced the TLR2 expression without an obvious change in the visceral fat volume in ob/ob mice. These results indicate that low-dose GH supplementation possibly inhibits the high-fat induced change of the adipocytes to TLR2/TNF-alpha co-expressing cells through the action of IGF-1.

Polychlorinated biphenyls alter the expression of endothelial nitric oxide synthase mRNA in human umbilical vein endothelial cells.

Polychlorinated biphenyls (PCBs) are a group of persistent pollutants that are detected in maternal serum and umbilical cord, suggesting that fetal exposure also needs to be considered. The effects of dioxin-like PCB congeners 3,3',4,4'-tetrachlorobiphenyl (PCB77) and 3,3',4,4',5-pentachlorobiphenyl (PCB126) and a non-dioxin-like compound 2,2',4,4',5,5'-hexachlorobiphenyl (PCB153) on the expression of endothelial nitric oxide synthase (eNOS), known to maintain blood flow to the fetus, in human umbilical vein endothelial cells (HUVECs) were investigated. The mRNA levels of eNOS, aryl hydrocarbon receptor (AhR) and cytochrome P450 (CYP) 1A1 in cells treated with 5 microM PCBs for 24 hours were analysed by real-time RT-PCR. Cells were also treated with alpha-naphthoflavone (alpha NF), an AhR antagonist or ICI 182780, an estrogen receptor (ER) antagonist, one hour prior to PCB exposure, to observe the effects of these receptors on eNOS modulation. Each PCB increased the eNOS mRNA level by 4.5-fold that was markedly inhibited by alphaNF. ERs were also suspected of altering eNOS levels because ICI 182780 treatment resulted in a decrease in the eNOS level. These results suggest that the eNOS mRNA expression increases due to the action of PCBs related to both AhR and ERs in HUVECs, and that maternal PCB exposure could influence fetal circulation.

CD151 dynamics in carcinoma-stroma interaction: integrin expression, adhesion strength and proteolytic activity.

A member of tetraspanin CD151 is a scaffold protein of laminin-binding integrins and it plays an important role in stable interaction between cells and basement membrane. Although the upregulation of CD151 in tumor cells is thought to accelerate tumor invasion and metastasis, detailed pathological investigation on CD151 and its association with integrins has not been well documented, yet. In the present study, we showed that the expression levels of CD151 and its associated integrin subunits in epidermal carcinoma cell HSC5 were higher than those in immortalized epidermal cell HaCaT. By the stimulation of epidermal growth factor, CD151 was dissociated from cell surface and dispersed in the cytoplasm, and alpha3beta1 integrin was concomitantly internalized. To understand the significance of CD151 in tumor cell dynamics, CD151 in HSC5 was knocked down (HSC5(CD151-)), and the expression of integrin subunits and matrix metalloproteinases (MMPs) were investigated. In HSC5(CD151-), striking morphological alteration on Matrigel and laminin, and cytoskeletal rearrangements were demonstrated. alpha3beta1 integrin was internalized in part, and alpha6beta4 integrin was re-distributed from basal site to cell periphery. Quantitative RT-PCR, Western blot and zymography revealed that the expression levels of MMP2, MMP7 and MMP9 were markedly downregulated in HSC5(CD151-). Immunoprecipitation assay demonstrated that MMP7 was co-immunoprecipitated with CD151. In double stainings, MMP7 was colocalized with CD151 at the leading edge of lamellipodia under migratory status. These results elucidated the importance of CD151 as one of the key molecules for integrin-dependent carcinoma-stroma interaction. It is indicated that CD151 might contribute not only to cell stabilization by associating with adhesion complexes but also to cell migration by inducing integrins re-localization and MMPs production.

A simple reconstruction for congenital unilateral lower lip palsy.

BACKGROUND: Congenital unilateral lower lip palsy or congenital hypoplasia of the depressor anguli oris muscle, also known as asymmetric crying facies, characterized by deformity of the lower lip, lacks aggressive surgical intervention methods. Although several dynamic and static reconstruction methods have been reported, textbooks introduce only passive surgical intervention, of weakening the unaffected side through techniques such as selective marginal mandibular neurectomy which, however, tends to produce lack of emotive movement. Therefore, a new surgical intervention for the reconstructive treatment of asymmetric crying facies is presented. METHODS: A bidirectional (horizontal and vertical) fascia was grafted to restore the aesthetic appearance of the asymmetric lower lip. The horizontal fascial strip achieves restoration of the center of the lower lip to its proper position, whereas the vertical fascial strip achieves aesthetic symmetry of the lower lip at mouth opening. Each end of the vertical strip is anchored to the lower lip and the mandibular bone, respectively, thereby allowing simultaneous movement of the mandible and lower lip. RESULTS: The reconstruction of unilateral lower lip palsy has been successfully performed on seven patients, with ages ranging from 2 years 9 months to 11 years 1 month, since June of 1996. CONCLUSIONS: The aim of this procedure is not to achieve complete dynamic reanimation. However, with regard to its simplicity and minimal invasiveness and the satisfaction of the patients, it is considered to be a well-balanced surgical intervention.

Time-course changes in the expression of heme oxygenase-1 in human subcutaneous hemorrhage.

To determine the time-course of human subcutaneous hemorrhage, heme oxygenase (HO)-1 expression and macrophage infiltration were observed using an immunohistochemical technique and semiquantitative analysis. The number of immunoreactive cells and the number of all infiltrating cells of each microscopic field were counted, and the ratio of the former to the latter was calculated as the positive cells ratio. An increase in the HO-1-positive cells ratio was observed starting at 3 h after injury, and the maximum ratio was observed 3 days after injury. The pattern of the increase in the macrophage ratio was similar to that of the HO-1-positive cells ratio in the early period after injury. Observation of serial sections revealed that the expression of HO-1 in the cells corresponded to the localization of macrophage. The present results suggest that the determination of HO-1 expression, as derived from macrophages, might be useful for the estimation of the time-course of subcutaneous hemorrhage.

Revascularization determines volume retention and gene expression by fat grafts in mice.

Autologous fat transplantation is a popular and useful technique in plastic and reconstructive surgery. The efficiency and survival of such grafts is predictable in many cases, but there are still issues to be resolved, such as how to improve graft volume retention. To address the issue of volume retention, we studied the effect of revascularization from the recipient on the size and function of adipocytes in fat grafts. Treatment of mice with TNP-470, an angiogenesis inhibitor, reduced blood flow from the recipient into the graft after subcutaneous transplantation of epididymal fat. The weight of transplanted tissues and the size of adipocytes in the grafts were significantly lower in mice treated with TNP-470 (TNP mice) than in control mice. Expression of genes for enzymes related to lipid accumulation was decreased in the grafts of TNP mice compared with control mice. Moreover, the expression of adipocyte-derived angiogenic peptides, VEGF and leptin, was significantly lower in the grafts of TNP mice than in grafts from control animals. The expression of VEGF and leptin by cultured human adipocytes was increased in the presence of conditioned medium from cultured vascular endothelial cells. These results show that the inhibition of the revascularization of fat grafts after transplantation reduces graft volume retention and cellular function. Early and adequate revascularization may be important for both the supply of nutrients and vasoactive interactions between vascular endothelial cells and adipocytes in graft.

Induction of uPA release in human peripheral blood lymphocytes by [deamino-Cysl,D-Arg8]-vasopressin (dDAVP).

[deamino-Cys(l),d-Arg(8)]-vasopressin (dDAVP), known to be an arginine vasopressin (AVP) V(2) receptor agonist, is an agent that increases fibrinolytic activity levels in plasma after its infusion into the human body. However, mechanisms underlying an increase and exact localization of the extrarenal dDAVP-responsive V(2) receptor remain unclarified. Two AVP receptors, V(1a) and V(2), and a related oxytocin (OT) receptor were found to be expressed in human lymphocytes. Furthermore, we found an increase of fibrinolytic activity in the medium of peripheral lymphocytes obtained from human volunteers less than 20 min after dDAVP infusion. The increased activity was also detected in the medium after incubating the lymphocytes in the presence of dDAVP in vitro, being highest at 20 min after the incubation. In accord with the increased fibrinolytic activity, the levels of urokinase-type plasminogen activator (uPA) in the medium were also increased. However, there was no significant difference of plasminogen activator inhibitor-1 (PAI-1), pro-uPA, and tissue-type plasminogen activator (tPA) concentrations in the medium between dDAVP treatment and control. When lymphocytes were preincubated with a V(2) receptor antagonist [Adamantaneacetyl(1),O-Et-d-Tyr(2),Val(4),Aminobutyryl(6),Arg(8,9)]-vasopressin, the dDAVP-induced uPA increase was diminished. In contrast, preincubation with a V(1) receptor antagonist, [beta-Mercapto-beta,beta-cyclopentamethylenepropionyl(1),O-Me-Tyr(2),Arg(8)]-vasopressin, prior to dDAVP treatment resulted in a greater increase of the uPA concentration in the medium than with the dDAVP treatment alone. Thus it was suggested that dDAVP may induce uPA release from human lymphocytes via V(2) receptor-mediated reaction, and also via cross-talk between V(1) and V(2) receptors.

Cyclic stretch induced MAPK phosphorylation stronger than static stretch.

The environment for living organism in space has microgravity and/or hypergravity and/or any kind of mechanical stresses. Cellular response may differ from the variety of mechanical stress. Mitogen-activated protein kinases (MAPKs) pathway is related to various cellular events. In the present study it was investigated the serial measurement of MAPK phosphorylation using western-blotting analysis following with three types of cyclic stretch, static, 0.1 Hz and 0.25 Hz. The result was that induction of MAPK phosphorylation had peaks within 2 to 4 hours and attenuated, while induction of p38 phosphorylation in 0.1 Hz stretch had a peak at 6 hours later and the strongest. Thus, there might be differential cellular response depends upon the frequency of cyclic stretch .

Increased expression of the Huntingtin interacting protein-1 gene in cells from Hutchinson Gilford Syndrome (Progeria) patients and aged donors.

Hutchinson Gilford syndrome (progeria [PG]) is a human disease associated with accelerated aging. To elucidate the acceleration mechanism, we first tried to transform a PG-derived cell line by infection of a recombinant adenovirus expressing HPV (human papilloma virus)-E6 and HPV-E7 genes. The transfected PG cells had a greater number of population doublings (PD) (>80), faster doubling time, and less staining of senescence-associated ss-galactosidase than the nontransfected PG cells. The transfected cells also showed markedly more detectable telomerase activity than the nontransformed cells. The expression levels of the genes in the E6-transduced and E7-transduced cell line were then compared with those of the nontransfected cell line using an mRNA differential display method, following reverse-transcriptase polymerase chain reaction analysis. Expression of huntingtin interacting protein-1 (HIP-1) gene was found to be increased not only in PG cells but also in fibroblast cells from aged healthy donors. Thus, HIP-1 might be a molecular assistant in the pathogenesis of the cellular senescent process in the human cells tested.

Increased levels of UV-induced protease activity in human UVAP-1 cells exposed to gravity-changing stress: involvement of E-64-sensitive proteases in suppression of UV mutagenicity.

Under the 1G condition, the increase in antipain-sensitive protease activity promptly after UV (mainly 254 nm wavelength) irradiation in cultured human cells is detected and found to be one of the intriguing events involved in suppression of cell mutability. It was found that two cell lines, RSa and its variant UVAP-1 cells are applicable; the former is hypermutable and not susceptible to protease activation, while the latter is hypomutable and susceptible. In the present study it was investigated whether the increase in protease activity by UV irradiation is also observed in hypomutable human UVAP-1 cells exposed to gravity-changing stress and whether the increase is involved in suppression of UV mutagenicity. Exposure of human UVAP-1 cells to gravity-changing stress such as free-fall and parabolic flight prior to UV irradiation resulted in a pronounced increase in protease activity, but not to hypergravity conditions (2 and 10G) prior to UV irradiation. To characterize the proteases, components of lysates from the cells exposed to free-fall prior to UV irradiation were fractionated by high performance liquid chromatography, indicating two separate fractions with highly increased levels of E-64-sensitive protease activity. In the cells treated with E-64 during their exposure to free-fall, K-ras codon 12 base substitution mutation was detected after UV irradiation, although the mutation was not detected after UV irradiation alone. Thus, the increase in E-64-sensitive protease activity may be involved in the suppression of UV mutagenicity in UVAP-1 cells exposed to free-fall.