Integrin α5 regulates motility of human monocyte-derived Langerhans cells during immune response.

Langerhans cells (LCs) are mainly present in the epidermis and mucosa, and have important roles during skin infection. Migration of LCs to lymph nodes is essential for antigen presentation. However, due to the difficulties in isolating and culturing human LCs, it is not fully understood how LCs move and interact with the extracellular matrix (ECM) through their adhesion molecules such as integrin, during the immune responses. In this study, we aimed to investigate LC motility, cell shape and the role of integrin under inflammatory conditions using monocyte-derived Langerhans cells (moLCs) as a model. As a result, lipopolysaccharide (LPS) stimulation increased adhesion on fibronectin coated substrate and integrin α5 expression in moLCs. Time-lapse imaging of moLCs revealed that stimulation with LPS elongated cell shape, whilst decreasing their motility. Additionally, this decrease in motility was not observed when pre-treated with a neutralising antibody targeting integrin α5. Together, our data suggested that activation of LCs decreases their motility by promoting integrin α5 expression to enhance their affinity to the fibronectin, which may contribute to their migration during inflammation.

Gap junction-mediated contraction of myoepithelial cells induces the peristaltic transport of sweat in human eccrine glands.

Eccrine sweat glands play an essential role in regulating body temperature. Sweat is produced in the coiled secretory portion of the gland, which is surrounded by obliquely aligned myoepithelial cells; the sweat is then peristaltically transported to the skin surface. Myoepithelial cells are contractile and have been implicated in sweat transport, but how myoepithelial cells contract and transport sweat remains unexplored. Here, we perform ex vivo live imaging of an isolated human eccrine gland and demonstrate that cholinergic stimulation induces dynamic contractile motion of the coiled secretory duct that is driven by gap junction-mediated contraction of myoepithelial cells. The contraction of the secretory duct occurs segmentally, and it is most prominent in the region surrounded by nerve fibers, followed by distension-contraction sequences of the excretory duct. Overall, our ex vivo live imaging approach provides evidence of the contractile function of myoepithelial cells in peristaltic sweat secretion from human eccrine glands.

Roles of TRPM4 in immune responses in keratinocytes and identification of a novel TRPM4-activating agent.

The skin is a protective interface between the internal organs and environment and functions not only as a physical barrier but also as an immune organ. However, the immune system in the skin is not fully understood. A member of the thermo-sensitive transient receptor potential (TRP) channel family, TRPM4, which acts as a regulatory receptor in immune cells, was recently reported to be expressed in human skin and keratinocytes. However, the function of TRPM4 in immune responses in keratinocytes has not been investigated. In this study, we found that treatment with BTP2, a known TRPM4 agonist, reduced cytokine production induced by tumor necrosis factor (TNF) α in normal human epidermal keratinocytes and in immortalized human epidermal keratinocytes (HaCaT cells). This cytokine-reducing effect was not observed in TRPM4-deficient HaCaT cells, indicating that TRPM4 contributed to the control of cytokine production in keratinocytes. Furthermore, we identified aluminum potassium sulfate, as a new TRPM4 activating agent. Aluminum potassium sulfate reduced Ca2+ influx by store-operated Ca2+ entry in human TRPM4-expressing HEK293T cells. We further confirmed that aluminum potassium sulfate evoked TRPM4-mediated currents, showing direct evidence for TRPM4 activation. Moreover, treatment with aluminum potassium sulfate reduced cytokine expression induced by TNFα in HaCaT cells. Taken together, our data suggested that TRPM4 may serve as a new target for the treatment of skin inflammatory reactions by suppressing the cytokine production in keratinocytes, and aluminum potassium sulfate is a useful ingredient to prevent undesirable skin inflammation through TRPM4 activation.

Basal cell carcinoma of the umbilicus associated with sweat gland structures: Two case reports.

RATIONALE: Basal cell carcinoma (BCC) arising in the umbilicus is relatively rare, and in particular, there have been few reports mentioning peritumoral sweat gland structures histopathologically. We herein, report 2 cases of umbilical BCC with sweat gland structures within and around the tumor. PATIENT CONCERNS: A 61-year-old woman had a 2-year history of black exudative plaque in her umbilicus, and an 80-year-old woman had a 6-month history of dark brownish plaque in the umbilicus, with exudation 2 months prior to her first visit. DIAGNOSES: Based on the histopathological finding, both cases were confirmed as BCC. The results of immunohistochemical staining showed that the tumor cells were Ber-EP4 positive. In addition, EMA-positive glandular structures were seen within and around the tumor. INTERVENTIONS: Curative resection at the level of the linea alba on the bottom side was performed. OUTCOMES: No relapse has been observed since resection in either patient. LESSONS: We herein report 2 cases of umbilical BCC with sweat glands and ducts. Although whether peri- and/or intra-tumor sweat gland structures are the source of the tumor or arise by transdifferentiation from tumor cells remains unclear, these findings may provide clues to help understand the morphopathogenesis of umbilical BCC in the future.

Anti-Inflammatory Role of TRPV4 in Human Macrophages.

The pathology of skin immune diseases such as atopic dermatitis is closely related to the overproduction of cytokines by macrophages. Although the pathological functions of macrophages in skin are known, mechanisms of how they detect the tissue environment remain unknown. TRPV4, a nonselective cation channel with high Ca2+ permeability, is activated at physiological temperatures from 27 to 35°C and involved in the functional control of macrophages. However, the relationship between TRPV4 function in macrophages and skin immune disease is unclear. In this study, we demonstrate that TRPV4 activation inhibits NF-κB signaling, resulting in the suppression of IL-1ß production in both human primary monocytes and macrophages derived from human primary monocytes. A TRPV4 activator also inhibited the differentiation of human primary monocytes into GM-CSF M1 macrophages but not M-CSF M2 macrophages. We also observed a significant increase in the number of inducible NO synthase-positive/TRPV4-negative dermal macrophages in atopic dermatitis compared with healthy human skin specimens. Our findings provide insight into the physiological relevance of TRPV4 to the regulation of macrophages during homeostasis maintenance and raise the potential for TRPV4 to be an anti-inflammatory target.

[Anxiety-related factors among elder-care facility employees regarding admission of tuberculosis patients after inpatient-treatment completion].

Objective　Tuberculosis (TB) patients are discharged after confirming their non-infective status. However, elder-care facilities often refuse to admit discharged TB patients. As no study has investigated anxiety among elder-care facility employees, we aimed to identify anxiety-associated factors among elder-care facility employees regarding the post-discharge admission of TB patients who have completed inpatient treatment.Methods　Among the 74 elder-care facilities under the jurisdiction of the Ibaraki Public Health Center in Osaka, Japan, (we excludes facilities that provided only daycare services), and invited all 3,213 employees of the remaining 70 facilities to participate in this questionnaire-based survey. Copies of an anonymous, self-administered questionnaire were mailed to the manager of each facility and were further distributed among employees. Responses were initially collected individually and subsequently directly collected from each facility by a public health nurse at the center. The questionnaire items included: the presence/absence of anxiety, resistance, and/or a feeling of difficulty about admitting TB patients who had completed inpatient treatment ("anxiety"), age, sex, occupation, years of work, total experience caring for TB patients, and knowledge of TB. The correlation between the presence/absence of anxiety and each item was analyzed using the chi-square test.Results　Completed questionnaires were obtained from 1,950 employees (response rate, 60.7%), of which 1,290 without missing data for relevant items were analyzed. Anxiety was present in 987 (76.5%) respondents. A significantly higher proportion of anxiety was observed in relation to the occupation (care workers and helpers), experience of caring for TB patients (respondents without such experience), and among employees who incorrectly answered questions on knowledge of TB, such as the infectiveness of TB patients after discharge, their management, and the risk of developing TB following infection.Conclusion　The study identified anxiety-associated factors among employees of elder-care facilities about admitting TB patients who had completed inpatient treatment for TB. Therefore, anxiety-mitigating environments may need to be established for such employees to facilitate the admission of discharged TB patients and their smooth return of patients to their pre-TB lives.

Visualization and characterization of spore morphogenesis in Paenibacillus polymyxa ATCC39564.

Paenibacillus polymyxa is a spore-forming Gram-positive bacterial species. Both its sporulation process and the spore properties are poorly understood. Here, we investigated sporulation in P. polymyxa ATCC39564. When cultured at 37℃ for 24 h in sporulation medium, more than 80% of the total cells in the culture were spores. Time-lapse imaging revealed that cellular morphological changes during sporulation of P. polymyxa were highly similar to those of B. subtilis. We demonstrated that genetic deletion of spo0A, sigE, sigF, sigG, or sigK, which are highly conserved transcriptional regulators in spore forming bacteria, abolished spore formation. In P. polymyxa, spo0A was required for cell growth in sporulation medium, as well as for the initiation of sporulation. The sigE and sigF mutants formed abnormal multiple asymmetric septa during the early stage of sporulation. The sigG and sigK mutants formed forespores in the sporangium, but they did not become mature. Moreover, fluorescence reporter analysis confirmed compartment-specific gene expression of spoIID and spoVFA in the mother cell and spoIIQ and sspF in the forespore. Transmission electron microscopy imaging revealed that P. polymyxa produces multilayered endospores but lacking a balloon-shaped exosporium. Our results indicate that spore morphogenesis is conserved between P. polymyxa and B. subtilis. However, P. polymyxa genomes lack many homologues encoding spore-coat proteins that are found in B. subtills, suggesting that there are differences in the spore coat composition and surface structure between P. polymyxa and B. subtilis.

Isolation and Culture of Primary Oral Keratinocytes from the Adult Mouse Palate.

For years, most studies involving keratinocytes have been conducted using human and mouse skin epidermal keratinocytes. Recently, oral keratinocytes have attracted attention because of their unique function and characteristics. They maintain the homeostasis of the oral epithelium and serve as resources for applications in regenerative therapies. However, in vitro studies that use oral primary keratinocytes from adult mice have been limited due to the lack of an efficient and well-established culture protocol. Here, oral primary keratinocytes were isolated from the palate tissues of adult mice and cultured in a commercial low-calcium medium supplemented with a chelexed-serum. Under these conditions, keratinocytes were maintained in a proliferative or stem cell-like state, and their differentiation was inhibited even after increased passages. Marker expression analysis showed that the cultured oral keratinocytes expressed the basal cell markers p63, K14, and α6-integrin and were negative for the differentiation marker K13 and the fibroblast marker PDGFRα. This method produced viable and culturable cells suitable for downstream applications in the study of oral epithelial stem cell functions in vitro.

Constitutive cell surface expression of ZZ domain for the easy preparation of yeast-based immunosorbents.

We describe a novel expression cassette that enables efficient and constitutive expression of the ZZ domain derived from Staphylococcus aureus protein A on the yeast cell surface to easily prepare yeast-based immunosorbents. Using this expression cassette containing the PGK1 promoter, a secretion signal derived from α-factor, and a Flo1-derived anchor protein, we successfully created a yeast-based immunosorbent for human serum albumin.

Metabolomic Alteration of Oral Keratinocytes and Fibroblasts in Hypoxia.

The oxygen concentration in normal human tissue under physiologic conditions is lower than the atmospheric oxygen concentration. The more hypoxic condition has been observed in the cells with wound healing and cancer. Somatic stem cells reside in a hypoxic microenvironment in vivo and prefer hypoxic culture conditions in vitro. Oral mucosa contains tissue-specific stem cells, which is an excellent tissue source for regenerative medicine. For clinical usage, maintaining the stem cell in cultured cells is important. We previously reported that hypoxic culture conditions maintained primary oral keratinocytes in an undifferentiated and quiescent state and enhanced their clonogenicity. However, the metabolic mechanism of these cells is unclear. Stem cell biological and pathological findings have shown that metabolic reprogramming is important in hypoxic culture conditions, but there has been no report on oral mucosal keratinocytes and fibroblasts. Herein, we conducted metabolomic analyses of oral mucosal keratinocytes and fibroblasts under hypoxic conditions. Hypoxic oral keratinocytes and fibroblasts showed a drastic change of metabolite concentrations in urea cycle metabolites and polyamine pathways. The changes of metabolic profiles in glycolysis and the pentose phosphate pathway under hypoxic conditions in the oral keratinocytes were consistent with those of other somatic stem cells. The metabolic profiles in oral fibroblasts showed only little changes in any pathway under hypoxia except for a significant increase in the antioxidant 2-oxoglutaric acid. This report firstly provides the holistic changes of various metabolic pathways of hypoxic cultured oral keratinocytes and fibroblasts.

Increase in primary cilia in the epidermis of patients with atopic dermatitis and psoriasis.

Primary cilia influence cell activity, and thus have a unique role in maintaining cell proliferation and differentiation. In atopic dermatitis (AD) and psoriasis, areas of skin inflammation exhibit dysregulated keratinocyte homeostasis. The role of primary cilia in these conditions remains unclear. The objectives of this study is to elucidate the incidence of primary cilia in skin inflammation and the potential mechanism underlying the dysregulation of keratinocytes. Primary cilia were observed using immunofluorescence staining. Normal skin samples were compared with skin samples from patients with AD or psoriasis in terms of cilia numbers and length. The effect of cytokine stimulation on ciliogenesis in keratinocytes was analysed using a primary keratinocyte culture. IFT88, an important ciliary intraflagellar protein, was blocked in Th2 and Th17 cytokines-stimulated keratinocytes. These effects were analysed with quantitative polymerase chain reaction and Western blot. Significant increases in ciliated cells were observed in AD and psoriasis skin samples compared with normal skin samples. The stimulation of keratinocytes using Th2 and Th17 cytokines modulated the formation of primary cilia. The amount of IFT88 in the primary cilia associated with the phosphorylation of JNK, but not p38, in keratinocytes stimulated with interleukin-13, 17A and 22. An increase of ciliated cells in the epidermis may impair keratinocyte differentiation under stress conditions caused by inflammation in both AD and psoriasis patients.

Distinct differences in hypoxic responses between human oral mucosa and skin fibroblasts in a 3D collagen matrix.

The differences between oral mucosa and skin wound healing involving hypoxic responses of fibroblasts are poorly elucidated. In this study, we aimed to study the different hypoxic responses between oral and skin fibroblasts embedded in a three-dimensional (3D) collagen matrix to address the early stage of wound healing. Primary oral mucosa fibroblasts (OMFs) obtained from the retromolar area and skin fibroblasts (SFs) obtained from the abdomen were cultured in the 3D 'floating model' under either 21%, 5% or 1% O2 for 2 days. Cell viability under hypoxia was higher in the OMFs than in the SFs. Collagen gel contraction was suppressed under hypoxic conditions in both fibroblasts, consistent with the reduction of alpha smooth muscle actin expression, except for SFs under 1% O2. Subsequently, their gene expression profiles between 21 and 1% O2 concentrations were compared via microarray technology, and the expression profiles of the extracellular matrix (ECM)-associated proteins, including matrix metalloproteinases and collagens, were evaluated. The OMFs were more susceptible to 1% O2, and more of their genes were downregulated than the SFs'. Although the production and expression levels of ECM-associated proteins in both fibroblasts diminished under hypoxia, those levels in OMFs were significantly higher than those in SFs. In the case of single origin OMFs and SFs, our findings suggest that OMFs possess a higher baseline production capacity of several ECM-associated proteins than SFs, except type III collagen. The intrinsic hypoxic responses of OMFs may be attributed to a more favourable wound healing in oral mucosa.

Rac1-dependent phagocytosis of apoptotic cells by oral squamous cell carcinoma cells: A possible driving force for tumor progression.

Apoptotic cell death frequently occurs in human cancer tissues including oral squamous cell carcinoma (SCC), wherein apoptotic tumor cells are phagocytosed not only by macrophages but also by neighboring tumor cells. We previously reported that the engulfment of apoptotic SCC cells by neighboring SCC cells frequently occurs at the invading front. Therefore, we hypothesized that the phagocytosis of these apoptotic cells by tumor cells contributes to disease progression. Herein, using cultured oral SCC cells, we aimed to confirm whether tumor cells actually phagocytose apoptotic cells and to examine whether cellular activities are regulated by the phagocytosis of apoptotic cells. Co-culture experiments showed that living cells could ingest apoptotic cells into phagolysosomes. NSC23766, an inhibitor of Rac1, which is a key regulator of phagocytic cup formation in professional phagocytes, dramatically suppressed the phagocytosis of apoptotic cells by living cells. Additionally, cell migration and the secretion of DKK1, a tumor-promoting protein, were enhanced by co-culture with apoptotic cells, whereas NSC23766 inhibited these effects. These results show that tumor cells can actively phagocytose apoptotic neighbors in a Rac1-dependent manner and that such activity increases their migration. The regulation of apoptotic cell phagocytosis thus represents new directions for therapeutic intervention for oral cancer.

ROCK inhibitors enhance bone healing by promoting osteoclastic and osteoblastic differentiation.

Osteoclast and osteoblast are essential for proper bone development and remodeling as well as recovery of bone fracture. In this study, we seek chemical compounds that enhance turnover of bone metabolism for promoting bone healing. First, we screen a chemical library which includes 378 compounds by using murine pre-osteoclastic RAW264.7 cells to identify compounds that promote osteoclastic differentiation. We find that two ROCK (Rho-associated coiled-coil kinase) inhibitors, HA-1077 (Fasudil) and Y-27632, enhance osteoclastogenesis. Subsequently, we identify that these two compounds also increase osteoblastic differentiation of MC3T3-E1 cells. Finally, our in vivo experiment shows that the local administration of ROCK inhibitors accelerate the bone healing of the rat calvarial defect.

Development of microstructured fish scale collagen scaffolds to manufacture a tissue-engineered oral mucosa equivalent.

The present study aimed to develop a more biomimetic tissue-engineered oral mucosa equivalent comprising 1% type I tilapia scale collagen scaffold having microstructures mimicking the dermal-epidermal junction of oral mucosa and oral keratinocytes as graft materials for human use. We designed four micropattern prototypes mimicking the dermal-epidermal junction. Using a semiconductor process and soft lithography, negative molds were fabricated to develop microstructures using both polydimethylsiloxane and silicon substrates. Micropattern configurations of dermal-epidermal junctions manufactured from fish collagen consisting of a fibril network using our micropatterning system were well preserved, although the internal fibril network of the pillar pattern was sparse. Mixing 1% chondroitin sulfate with the collagen matrix minimized tissue-engineered oral mucosa equivalent contraction. Histologic examinations showed a flattening of the vertical dimensions of all microstructures and expansion of their pitches, indicating changes in the originally designed configurations. Nonetheless, histologic examinations revealed that a fully differentiated and stratified epithelial layer was developed on all scaffolds, suggesting that the microstructured fish scale collagen scaffolds have potential in the manufacturing of tissue-engineered oral mucosa equivalents for clinical use; however, enhancement of the mechanical properties of micropatterns is required. Our micropatterning technology can also apply to the development of oral mucosa in vitro models.

A Questionnaire Survey of the Inhalation Instruction in Pharmacies.

Few studies have focused on the inhalation instruction in pharmacies which have the crucial role on the inhalation instruction. The aim of this study is to evaluate the level of knowledge and the degree of interest for asthma inhalation instruction methods among pharmacists receiving prescription from clinics. We conducted questionnaire surveys to chief pharmacists of 39 consecutive pharmacies belonging to HANSHIN Dispensing Pharmacy in Hyogo, Japan at July 2011. We obtained valid responses from 35 pharmacies. Among them, 14 pharmacies dealt with prescriptions mainly from the clinics (clinic pharmacies) and 21 pharmacies dealt with prescriptions originated from hospitals (hospital pharmacies), including 13 pharmacies that dealt with prescription filled by respiratory physicians (specialty hospital pharmacies). Although the inhalation instruction at the first visit was provided at every pharmacy, only 54.3% of all pharmacies provided inhalation instructions after the second visit. Compared to 0% of the clinic pharmacies, 40% of the specialty hospital pharmacies visually checked the patient's inhalation procedure after the second visit. Visual confirmation of the inhalation technique, especially in the clinic pharmacies, might play an important role in maintaining treatment adherence.

5-Aminolevulinic acid fermentation using engineered Saccharomyces cerevisiae.

BACKGROUND: 5'-Aminolevulinic acid (ALA) is widely used in the pharmaceutical industry, healthcare, and food production, and is a substrate for the biosynthesis of heme, which is required for respiration and photosynthesis. Enhancement of ALA biosynthesis has never been developed in Saccharomyces cerevisiae, which is a well-known model microorganism used for bioproduction of many value-added compounds. RESULTS: We demonstrated that metabolic engineering significantly improved ALA production in S. cerevisiae. First, we found that overexpression of HEM1, which encodes ALA synthetase, increased ALA production. Furthermore, addition of an optimal amount of glycine, a substrate for ALA biosynthesis, or levulinic acid, an inhibitor of ALA dehydrogenase, effectively increased ALA production. Next, we developed an assay for multiple metabolites including ALA and found that aconitase, encoded by ACO1 and ACO2, is the rate-limiting enzyme of ALA biosynthesis when sufficient glycine is supplied. Overexpression of ACO2 further enhanced ALA production in S. cerevisiae overexpressing HEM1. CONCLUSIONS: In this study, ALA production in S. cerevisiae was enhanced by metabolic engineering. This study also shows a strategy to identify the rate-limiting step of a target synthetic pathway by assay for multiple metabolites alongside the target product. This strategy can be applied to improve production of other valuable products in the well-studied and well-industrialized microorganism S. cerevisiae.

Noninvasive measurement of cell/colony motion using image analysis methods to evaluate the proliferative capacity of oral keratinocytes as a tool for quality control in regenerative medicine.

Image-based cell/colony analyses offer promising solutions to compensate for the lack of quality control (QC) tools for noninvasive monitoring of cultured cells, a regulatory challenge in regenerative medicine. Here, the feasibility of two image analysis algorithms, optical flow and normalised cross-correlation, to noninvasively measure cell/colony motion in human primary oral keratinocytes for screening the proliferative capacity of cells in the early phases of cell culture were examined. We applied our software to movies converted from 96 consecutive time-lapse phase-contrast images of an oral keratinocyte culture. After segmenting the growing colonies, two indices were calculated based on each algorithm. The correlation between each index of the colonies and their proliferative capacity was evaluated. The software was able to assess cell/colony motion noninvasively, and each index reflected the observed cell kinetics. A positive linear correlation was found between cell/colony motion and proliferative capacity, indicating that both algorithms are potential tools for QC.

Evaluation of the effectiveness of caspofungin against febrile neutropenia and the factors related to the alteration in its plasma concentration.

Caspofungin (CPFG) is an echinocandin antifungal agent that inhibits the synthesis of ß-1, 3-D-glucan, a critical component of the cell wall of target fungi. Several clinical studies have confirmed the efficacy and safety of CPFG in patients with febrile neutropenia (FN); however, there are no reports available in Japanese patients with FN. Therefore, we investigated the therapeutic efficacy and pharmacokinetics of CPFG as an empirical therapy in a Japanese hospital. Twenty-four Japanese patients, who were diagnosed with FN at Gifu University Hospital from February 2014 to August 2017, were enrolled. Blood samples were collected at the end of CPFG dosing (0.5 h after the infusion) on day 1 and immediately prior to the next infusion on days 2, 3, and 4. The concentration of CPFG in plasma was measured by high-performance liquid chromatography. The efficacy was assessed by five of the component endpoints, and safety was monitored according to the Common Terminology Criteria for Adverse Events. CPFG showed an excellent effect against FN (75%, 18/24), without any serious hepatic or renal toxicity. Regarding the pharmacokinetics, the plasma concentration of CPFG was significantly correlated with body weight; although, no correlation was observed between the plasma concentration of CPFG and the other factors investigated, such as gender or laboratory results. These results suggest the high efficacy, safety, and tolerability of CPFG as an empirical antifungal therapy for Japanese patients with FN.

Autopsied case of sporadic Creutzfeldt-Jakob disease classified as MM1+2C-type.

We encountered an autopsy case of sporadic Creutzfeldt-Jakob disease (CJD) pathologically classified as MM1+2C-type, where Western blot analysis of prion protein (PrP) mainly showed type-1 scrapie PrP (PrPSc ) but also, partially, mixed type-2 PrPSc . A Japanese woman complained of visual disorder at the age of 86 years and then showed disorientation and memory disturbances. Magnetic resonance imaging (MRI) showed cerebral cortical hyperintensity on diffusion-weighted images. The patient died 2 months after the onset of symptoms; her condition did not reach the akinetic mutism state and periodic sharp-wave complexes on electroencephalography and myoclonus were not recognized. The brain weighed 1100 g and neuropathological examination showed extensive fine vacuole-type spongiform changes in the cerebral cortex. In some cortical regions, large confluent vacuole-type spongiform changes were also present. Gliosis and hypertrophic astrocytosis were generally mild, and tissue rarefaction of the neuropil and neuronal loss were not apparent. PrP immunostaining showed diffuse synaptic-type PrP deposition in the cerebral gray matter, but some regions with large confluent vacuoles showed perivacuolar-type deposition. We speculated, based on the clinicopathological findings and previous reports, that most MM1-type sporadic CJD cases may be associated with type-2 PrPSc , at least partially, within certain regions of the cerebrum.