Analysis of composition and microstructure of diatom frustules in mud on the coast of Boryeong- city, South Korea.

The microstructure of diatom frustules found in mud sediments along the coast of Boryeong- city, South Korea, was observed using a scanning electron microscopy (SEM), and the constituent elements of diatoms were analyzed using energy-dispersive X-ray spectroscopy (EDS). Diatom frustules and clay minerals were present in the SEM images of the mud powder. High-magnification SEM images revealed that the surface of the frustules contained identically shaped circular pores, measuring 1 µm in diameter, arranged at regular intervals. This study revealed that the diatom shell fragments in the mud powder ranged in size from 3 to 30 µm, with an average thickness of approximately 2.5 µm. The elements Si, Al, Fe, K, Na, Mg, and Ti were detected while analyzing the frustule constituents, with Si being the primary component with the highest content.

Analysis of microstructural characteristics and components of red and yellow ink pigments used in permanent makeup.

Our purpose in this study is to analyze the microstructural characteristics and constituent elements of inorganic substances added to the yellow ink and red ink pigments used in permanent makeup. We observed the microstructural properties of inorganic pigments added to the ink using a scanning electron microscopy (SEM) and analyzed the constituent elements of the inorganic pigment particles using an energy dispersive X-ray spectroscopy (EDX). In red wine-colored ink, cubic titanium dioxide with a diameter of 110 to 200 nm was the major component, and rod-shaped iron oxide was rarely observed. Most of the ingredients of taupe yellow ink were rod-shaped yellow iron oxide, and a small amount of cubic titanium dioxide was observed. Red wine-colored ink and taupe yellow ink contained lumps composed of titanium dioxide particles. In red wine-colored ink, lumps were formed by agglomeration. However, we observed that the surface of the lump composed of titanium dioxide in the taupe yellow ink had a smooth surface caused by external physical compression. The titanium dioxide particle mass which found in taupe yellow ink in this study is an artificial product. When this mass accumulates in the dermis, it may cause a color mismatch. Therefore, permanent makeup using fine pigments should be free of foreign substances that may cause trouble in the skin. In addition, there is a need to improve the quality of the ink so that the required color can be safe and long lasting in the dermis.

Ultrastructure of the fertilized egg envelopes in Hemigrammus erythrozonus, Characidae, Teleostei.

We examined the morphology and ultrastructures of fertilized egg envelopes of glowlight tetra (Hemigrammus erythrozonus) belong to Characidae using light and electron microscopes.The fertilized eggs were spherical, transparent, demersal, adhesive, and have no oil droplet. The perivitelline space was well-developed and the micropyle was surrounded by 15-20 uplifted lines of egg envelope in a spoke like pattern. The outer surface of egg envelope was rough side with grooves. Also, the total thickness of the fertilized egg envelope was about 2.1-2.3 µm, and the fertilized egg envelope consisted of two layers, an outer adhesive electron-dense layer with grooves and three feather-like lamellae layers. Collectively, these morphological characteristics of fertilized egg and micropyle with spoke-like structure showed family Characidae specificity, and ultrastructures of outer surface and section of fertilized egg envelope showed species specificity.

IKKß-mediated inflammatory myeloid cell activation exacerbates experimental autoimmune encephalomyelitis by potentiating Th1/Th17 cell activation and compromising blood brain barrier.

BACKGROUND: The inflammatory myeloid cell activation is one of the hallmarks of experimental autoimmune encephalomyelitis (EAE), yet the in vivo role of the inflammatory myeloid cell activation in EAE has not been clearly resolved. It is well-known that IKK/NF-κB is a key signaling pathway that regulates inflammatory myeloid activation. METHODS: We investigated the in vivo role of inflammatory myeloid cell activation in myelin oligodendrocyte glycoprotein (MOG) peptides-induced EAE using myeloid cell type-specific ikkß gene conditional knockout-mice (LysM-Cre/Ikkß (F/F) ). RESULTS: In our study, LysM-Cre/Ikkß (F/F) mice had alleviated clinical signs of EAE corresponding to the decreased spinal demyelination, microglial activation, and immune cell infiltration in the spinal cord, compared to the wild-type mice (WT, Ikkß (F/F) ). Myeloid ikkß gene deletion significantly reduced the percentage of CD4(+)/IFN-γ(+) (Th1) and CD4(+)/IL-17(+) (Th17) cells but increased the percentages of CD4(+)/CD25(+)/Foxp3(+) (Treg) cells in the spinal cord and lymph nodes, corresponding to the altered mRNA expression of IFN-γ, IL-17, IL-23, and Foxp3 in the spinal cords of LysM-Cre/Ikkß (F/F) EAE mice. Also, the beneficial effect of myeloid IKKß deletion in EAE corresponded to the decreased permeability of the blood brain barrier (BBB). CONCLUSIONS: Our findings strongly suggest that IKK/NF-kB-induced myeloid cell activation exacerbates EAE by activating Th1 and Th17 responses and compromising the BBB. The development of NF-κB inhibitory agents with high efficacy through specific targeting of IKKß in myeloid cells might be of therapeutic potential in MS and other autoimmune disorders.

Korean Red Ginseng and Ginsenoside-Rb1/-Rg1 Alleviate Experimental Autoimmune Encephalomyelitis by Suppressing Th1 and Th17 Cells and Upregulating Regulatory T Cells.

The effects of Korean red ginseng extract (KRGE) on autoimmune disorders of the nervous system are not clear. We investigated whether KRGE has a beneficial effect on acute and chronic experimental autoimmune encephalomyelitis (EAE). Pretreatment (daily from 10 days before immunization with myelin basic protein peptide) with KRGE significantly attenuated clinical signs and loss of body weight and was associated with the suppression of spinal demyelination and glial activation in acute EAE rats, while onset treatment (daily after the appearance of clinical symptoms) did not. The suppressive effect of KRGE corresponded to the messenger RNA (mRNA) expression of proinflammatory cytokines (tumor necrosis factor-α [TNF-α] and interleukin [IL]-1ß), chemokines (RANTES, monocyte chemotactic protein-1 [MCP-1], and macrophage inflammatory protein-1α [MIP-1α]), adhesion molecules (intercellular adhesion molecule-1 [ICAM-1], vascular cell adhesion molecule-1 [VCAM-1], and platelet endothelial cell adhesion molecule [PECAM-1]), and inducible nitric oxide synthase in the spinal cord after immunization. Interestingly, in acute EAE rats, pretreatment with KRGE significantly reduced the population of CD4(+), CD4(+)/IFN-γ(+), and CD4(+)/IL-17(+) T cells in the spinal cord and lymph nodes, corresponding to the downregulation of mRNA expression of IFN-γ, IL-17, and IL-23 in the spinal cord. On the other hand, KRGE pretreatment increased the population of CD4(+)/Foxp3(+) T cells in the spinal cord and lymph nodes of these rats, corresponding to the upregulation of mRNA expression of Foxp3 in the spinal cord. Interestingly, intrathecal pretreatment of rats with ginsenosides (Rg1 and Rb1) significantly decreased behavioral impairment. These results strongly indicate that KRGE has a beneficial effect on the development and progression of EAE by suppressing T helper 1 (Th1) and Th17 T cells and upregulating regulatory T cells. Additionally, pre- and onset treatment with KRGE alleviated neurological impairment of myelin oligodendrocyte glycoprotein(35-55)-induced mouse model of chronic EAE. These results warrant further investigation of KRGE as preventive or therapeutic strategies for autoimmune disorders, such as multiple sclerosis.

Increased Expression of Osteopontin in the Degenerating Striatum of Rats Treated with Mitochondrial Toxin 3-Nitropropionic Acid: A Light and Electron Microscopy Study.

The mycotoxin 3-nitropropionic acid (3NP) is an irreversible inhibitor that induces neuronal damage by inhibiting mitochondrial complex II. Neurodegeneration induced by 3NP, which is preferentially induced in the striatum, is caused by an excess influx and accumulation of calcium in mitochondria. Osteopontin (OPN) is a glycosylated phosphoprotein and plays a role in the regulation of calcium precipitation in the injured brain. The present study was designed to examine whether induction of OPN protein is implicated in the pathogenesis of 3NP-induced striatal neurodegeneration. We observed overlapping regional expression of OPN, the neurodegeneration marker Fluoro-Jade B, and the microglial marker ionized calcium-binding adaptor molecule 1 (Iba1) in the 3NP-lesioned striatum. OPN expression was closely associated with the mitochondrial marker NADH dehydrogenase (ubiquinone) flavoprotein 2 in the damaged striatum. In addition, immunoelectron microscopy demonstrated that OPN protein was specifically localized to the inner membrane and matrix of the mitochondria in degenerating striatal neurons, and cell fragments containing OPN-labeled mitochondria were also present within activated brain macrophages. Thus, our study revealed that OPN expression is associated with mitochondrial dysfunction produced by 3NP-induced alteration of mitochondrial calcium homeostasis, suggesting that OPN is involved in the pathogenesis of striatal degeneration by 3NP administration.

Comparison of Alcian Blue, Trypan Blue, and Toluidine Blue for Visualization of the Primo Vascular System Floating in Lymph Ducts.

The primo vascular system (PVS), floating in lymph ducts, was too transparent to be observed by using a stereomicroscope. It was only detectable with the aid of staining dyes, for instance, Alcian blue, which was injected into the lymph nodes. Some dyes were absorbed preferentially by the PVS than the lymph wall. It remains a standing problem to know what dyes are absorbed better by the PVS than the lymph walls. Such information would be useful to unravel the biochemical properties of the PVS that are badly in need for obtaining large amount of PVS specimens. In the current work we tried two other familiar dyes which were used in PVS research before. We found that Trypan blue and toluidine blue did not visualize the PVS. Trypan blue was cleared by the natural washing. Toluidine blue did not stain the PVS, but it did leave stained spots in the lymph wall and its surrounding tissues, and it leaked out of the lymph wall to stain surrounding connective tissues. These completely different behaviors of the three dyes were found for the first time in the current work and provide valuable information to elucidate the mechanism through which some special dyes stained the PVS preferentially compared to the lymphatic wall.

Internalization of silver nanoparticles into mouse spermatozoa results in poor fertilization and compromised embryo development.

Silver nanoparticles (AgNPs) have many features that make them attractive as medical devices, especially in therapeutic agents and drug delivery systems. Here we have introduced AgNPs into mouse spermatozoa and then determined the cytotoxic effects of AgNPs on sperm function and subsequent embryo development. Scanning electron microscopy and transmission electron microscopy analyses showed that AgNPs could be internalized into sperm cells. Furthermore, exposure to AgNPs inhibited sperm viability and the acrosome reaction in a dose-dependent manner, whereas sperm mitochondrial copy numbers, morphological abnormalities, and mortality due to reactive oxygen species were significantly increased. Likewise, sperm abnormalities due to AgNPs internalization significantly decreased the rate of oocyte fertilization and blastocyst formation. Blastocysts obtained from AgNPs-treated spermatozoa showed lower expression of trophectoderm-associated and pluripotent marker genes. Overall, we propose that AgNPs internalization into spermatozoa may alter sperm physiology, leading to poor fertilization and embryonic development. Such AgNPs-induced reprotoxicity may be a valuable tool as models for testing the safety and applicability of medical devices using AgNPs.

Collagen complexes increase the efficiency of iPS cells generated using fibroblasts from adult mice.

Different interventions are being tested for restoration of the youthfulness of adult mouse-derived fibroblasts. However, fundamental issues, such as the decline of adult mouse-derived fibroblast activity with age, remain unresolved. Therefore, in this study, we examined whether treatment with collagen complexes has beneficial effects on the rejuvenation or reprogramming of adult mouse-derived fibroblasts. Further, we investigated the mechanisms of rejuvenation of adult mouse-derived fibroblasts during treatment with total collagen complexes. We isolated total collagen complexes from the tails of young mice and cultured adult mouse-derived fibroblasts with or without the collagen complexes. When compared with fibroblasts cultured without collagen complexes, adult-derived fibroblasts cultured with collagen complexes over five consecutive passages showed a more youthful state, expanded at a higher rate, and exhibited reduced spontaneous cell death. The fibroblasts cultured in the presence of collagen complexes also showed extensive demethylation in the promoter regions of cell cycle-related genes such as PCNA, increased proliferation, and decreased senescence. In addition, the efficiency of reprogramming of fibroblasts to become induced pluripotent stem (iPS) cells was significantly higher in young- and adult-derived fibroblasts cultured with collagen complexes than in adult-derived fibroblasts cultured alone. Furthermore, mechanistic evidence shows that genes involved in anti-proliferative pathways, including Ink4a/Arf locus genes and p53, were downregulated in fibroblasts exposed to collagen complexes. Interestingly, our results suggest that the rejuvenation process was mediated via the α2ß1 integrin-dependent Bmi-1 pathway. Thus, collagen complexes both stimulate proliferation and inhibit cell death and growth arrest in fibroblasts, which appears to be a promising approach for improving the efficiency of reprogramming.

Observation of the Primo Vessel Approaching the Axillary Lymph Node with the Fluorescent Dye, DiI.

The primo vascular system (PVS) floating in lymph fluid has mostly been observed in large caliber ducts around the caudal vena cava and the thoracic duct of rabbits, rats, and mice. But the PVS has not been traced up to the lymph nodes. It has not been established whether the PVS leaves the lymph vessel through the lymph vessel wall or it enters the lymph nodes. Therefore, observing the PVS entering a lymph node, for example, the axillary node, is desirable. In the current work, we traced the PVS approaching up to the surface of axillary node of a rat. The method used for this study was based upon a method that was recently developed to detect the PVS in the lymph duct from the inguinal to the axillary nodes in the skin of a rat by injecting Alcian blue into the inguinal node. However, the Alcian blue blurred near the lymph nodes and tracing the PVS up to the lymph nodes has not been possible. The current method clearly showed the PVS approaching the axillary node.

MicroRNA dysregulation in liver and pancreas of CMP-Neu5Ac hydroxylase null mice disrupts insulin/PI3K-AKT signaling.

CMP-Neu5Ac hydroxylase (Cmah)-null mice fed with a high-fat diet develop fasting hyperglycemia, glucose intolerance, and pancreatic ß-cell dysfunction and ultimately develop characteristics of type 2 diabetes. The precise metabolic role of the Cmah gene remains poorly understood. This study was designed to investigate the molecular mechanisms through which microRNAs (miRNAs) regulate type 2 diabetes. Expression profiles of miRNAs in Cmah-null mouse livers were compared to those of control mouse livers. Liver miFinder miRNA PCR arrays (n = 6) showed that eight miRNA genes were differentially expressed between the two groups. Compared with controls, seven miRNAs were upregulated and one miRNA was downregulated in Cmah-null mice. Specifically, miR-155-5p, miR-425-5p, miR-15a-5p, miR-503-5p, miR-16-5p, miR-29a-3p, and miR-29b-3p were significantly upregulated in the liver and pancreas of Cmah-null mice. These target miRNAs are closely associated with dysregulation of insulin/PI3K-AKT signaling, suggesting that the Cmah-null mice could be a useful model for studying diabetes.

Gene expression and pathway analysis of effects of the CMAH deactivation on mouse lung, kidney and heart.

BACKGROUND: N-glycolylneuraminic acid (Neu5Gc) is generated by hydroxylation of CMP-Neu5Ac to CMP-Neu5Gc, catalyzed by CMP-Neu5Ac hydroxylase (CMAH). However, humans lack this common mammalian cell surface molecule, Neu5Gc, due to inactivation of the CMAH gene during evolution. CMAH is one of several human-specific genes whose function has been lost by disruption or deletion of the coding frame. It has been suggested that CMAH inactivation has resulted in biochemical or physiological characteristics that have resulted in human-specific diseases. METHODOLOGY/PRINCIPAL FINDINGS: To identify differential gene expression profiles associated with the loss of Neu5Gc expression, we performed microarray analysis using Illumina MouseRef-8 v2 Expression BeadChip, using the main tissues (lung, kidney, and heart) from control mice and CMP-Neu5Ac hydroxylase (Cmah) gene knock-out mice, respectively. Out of a total of 25,697 genes, 204, 162, and 147 genes were found to be significantly modulated in the lung, kidney, and heart tissues of the Cmah null mouse, respectively. In this study, we examined the gene expression profiles, using three commercial pathway analysis software packages: Ingenuity Pathways Analysis, Kyoto Encyclopedia of Genes and Genomes analysis, and Pathway Studio. The gene ontology analysis revealed that the top 6 biological processes of these genes included protein metabolism and modification, signal transduction, lipid, fatty acid, and steroid metabolism, nucleoside, nucleotide and nucleic acid metabolism, immunity and defense, and carbohydrate metabolism. Gene interaction network analysis showed a common network that was common to the different tissues of the Cmah null mouse. However, the expression of most sialytransferase mRNAs of Hanganutziu-Deicher antigen, sialy-Tn antigen, Forssman antigen, and Tn antigen was significantly down-regulated in the liver tissue of Cmah null mice. CONCLUSIONS/SIGNIFICANCE: Mice bearing a human-like deletion of the Cmah gene serve as an important model for the study of abnormal pathogenesis and/or metabolism caused by the evolutionary loss of Neu5Gc synthesis in humans.

Histological study on the effect of electrolyzed reduced water-bathing on UVB radiation-induced skin injury in hairless mice.

Electrolyzed reduced water (ERW), functional water, has various beneficial effects via antioxidant mechanism in vivo and in vitro. However there is no study about beneficial effects of ERW bathing. This study aimed to determine the effect of ERW bathing on the UVB-induced skin injury in hairless mice. For this purpose, mice were irradiated with UVB to cause skin injury, followed by individually taken a bath in ERW (ERW-bathing) and tap water (TW-bathing) for 21 d. We examined cytokines profile in acute period, and histological and ultrastructural observation of skin in chronic period. We found that UVB-mediated skin injury of ERW-bathing group was significantly low compared to TW control group in the early stage of experiment. Consistently, epidermal thickening as well as the number of dermal mast cell was significantly lowered in ERW-bathing group. Defection of corneocytes under the scanning electron microscope was less observed in ERW-bathing group than in TW-bathing group. Further, the level of interleukin (IL)-1ß, tumor necrosis factor (TNF)-α and IL-12p70 in ERW group decreased whereas those of IL-10 increased. Collectively, our data indicate that ERW-bathing significantly reduces UVB-induced skin damage through influencing pro-/anti-inflammatory cytokine balance in hairless mice. This suggests that ERW-bathing has a positive effect on acute UVB-mediated skin disorders. This is the first report on bathing effects of ERW in UVB-induced skin injury.

The immunological effects of electrolyzed reduced water on the Echinostoma hortense infection in C57BL/6 mice.

Electrolyzed reduced water (ERW) is widely used for drinking by people in Asia. The purpose of this study was to examine the immunological effect of ERW on the immunity of animals by supplying ERW to C57BL/6 mice infected with Echinostoma hortense metacercariae. In the non-infected groups, interleukin (IL)-4 (p < 0.001), IL-5, IL-10, IL-1beta, tumor necrosis factor (TNF)-alpha and immunoglobulin (Ig) A expression of the group fed ERW (ERW group) increased in small intestine compared with the normal control group. In the case of infected groups, the group fed ERW (ERW+E. hortense group) showed the result that IL-4, IL-5, IL-10 and Ig A expression increased, but IL-1beta and TNF-alpha (p < 0.001) decreased, and the number of goblet cells (p < 0.001) and helix pomatia agglutinin (HPA) positive cells increased compared with the group without feeding ERW. However, adult worm recovery rate was markedly increased (p < 0.05). On the other hand, the expression of all the cytokines except IL-10 in spleen was mildly increased but not significant statistically, and there was no significant difference in the numerical changes of white blood cell (WBC). These results indicate that feeding ERW may have influence on the local immune response (Th-1 type cytokines such as IL-1beta, TNF-alpha) in the small intestine but not on the systemic immune response.

Three-dimensional reconstruction of medieval child mummy in Yangju, Korea, using multi-detector computed tomography.

Recently reported studies on the medieval mummies in Korea have been regarded as an invaluable source for studying the physical characteristics of medieval Koreans. However, since the reported medieval mummies were re-buried by their descendants without any scientific investigations, the development of a brief, non-invasive investigation technique was desperately needed among the researchers in Korea. In this regard, we tried to apply high-quality multi-detector computed tomography (MDCT) with three-dimensional (3D) reconstructions and multi-planar reformat (MPR) to investigate Korean mummies. In our study, after 1.25 or 2.5mm thin slice axial images were taken, 3D reconstruction and MPR were performed to get more accurate information about internal organs. In this trial, we successfully showed high-quality images for the brain, muscles, bones, heart and liver. During various trials for getting selected organs, we could make the 3D reconstructed images of them. Since we could show that the current MDCT technique could be useful for obtaining high-quality 3D reconstructed images of the internal organs of Korean mummies, this technique will be used in forthcoming similar cases, which could not be investigated using invasive techniques.

Electron microscopic study of novel threadlike structures on the surfaces of mammalian organs.

The ultrastructures of novel threadlike structures (NTSs) and corpuscles on the surfaces of internal organs of rats were investigated using electron microscopy. The samples were studied in situ by using a stereomicroscope and were taken for further morphological analysis. Scanning electron microscope (SEM) images revealed a bundle structure of threadlike tissue, which was composed of several 10-micro m-thick subducts. The surfaces of the corpuscles were rather coarse and fenestrated. The corpuscles had cucumber-like shapes with an average length of about 2 mm and a thickness of about 400 micro m. Transmission electron microscope (TEM) images disclosed disordered collagen fibers, which formed the extracellular matrix of the threadlike tissue, and immune-function cells, like macrophages, mast cells, and eosinophils. Sinuses of various diameters, which were thought to be cross-sections of the lumens of the subducts, were observed in the TEM, cryo-SEM and focused-ion-beam SEM images. These SEM images were obtained for the first time to reveal the detailed structure of the NTSs that were only recently discovered.

Preserved skin structure of a recently found fifteenth-century mummy in Daejeon, Korea.

Recently published reports on Korea's medieval mummies have been regarded as an invaluable source for studies into the physical characteristics of medieval Koreans. However, even though the mummified tissues have been investigated histologically on various previous occasions, there are many unanswered questions relating to their tissue preservation. The aim of this study was to obtain new data on the ultramicroscopic characteristics of the mummified skin of a fifteenth-century mummy found recently in Daejeon--one of the oldest ever found in Korea. Electron microscopy revealed that much of the epidermis had decayed; what remained of the dermis was filled with collagen fibres and melanin granules or invading bacterial spores present within the mummified epidermis. Considering the histological characteristics shared by naturally formed mummies in different parts of the world, we concluded that the ultramicroscopic patterns of the Daejeon mummy were more comparable with those naturally formed mummies than with artificially formed ones. This is the first full description of the morphological characteristics of the skin collected from this recently found medieval mummy from Daejeon, South Korea.

Endoscopic investigation of the internal organs of a 15th-century child mummy from Yangju, Korea.

Our previous reports on medieval mummies in Korea have provided information on their preservation status. Because invasive techniques cannot easily be applied when investigating such mummies, the need for non-invasive techniques incurring minimal damage has increased among researchers. Therefore, we wished to confirm whether endoscopy, which has been used in non-invasive and minimally invasive studies of mummies around the world, is an effective tool for study of Korean mummies as well. In conducting an endoscopic investigation on a 15th-century child mummy, we found that well-preserved internal organs remained within the thoracic, abdominal and cranial cavities. The internal organs - including the brain, spinal cord, lung, muscles, liver, heart, intestine, diaphragm and mesentery - were easily investigated by endoscopy. Even the stool of the mummy, which accidentally leaked into the abdominal cavity during an endoscopic biopsy, was clearly observed. In addition, unusual nodules were found on the surface of the intestines and liver. Our current study therefore showed that endoscopic observation could provide an invaluable tool for the palaeo-pathological study of Korean mummies. This technique will continue to be used in the study of medieval mummy cases in the future.

Ultramicroscopic study on the hair of newly found 15th century mummy in Daejeon, Korea.

Recent reports on the medieval mummies in Korea have been an invaluable source for the studies on the physical characteristics of medieval Koreans. Though the histological observations on the medieval mummy found in Korea were actively reported by researchers, it could not be said that all the queries on the mummies had been completely answered at the present time. In this study, we tried to add the novel data on the ultramicroscopic characteristics of the hair taken from the recently discovered 15th century mummy, the oldest one ever found in Korea. Even comparing with the hair from living individual, the hair of 15th century mummy showed very intact appearances during observation with electron microscope because the scales on the surface of the mummified hair were not damaged, which were not easily maintained even in living individuals. As to the cause of the excellent preservation of 15th century mummified hair, the presence of surface coat on the hair should be considered. That is, just like the complete sealing effect of lime-soil mixture barrier around the coffin, the presence of the surface coat including calcium and sodium might inhibit the water or fungal infiltrations into the hair shaft.

Ultramicroscopic observations on morphological changes in hair during 25 years of weathering.

Weathering or long-term burial may cause profound morphological and histological changes in hair, which may affect the results of forensic and archaeological investigations. We therefore used ultramicroscopic techniques to assay the changes in weathering hair shafts caused by burial for up to 25 years. We found that the middle portion of hair shafts from living individuals shows the expected histological hair structure, while the cuticle layer was absent from the terminal portion of the same hairs, which may be due to the increased weathering experienced by the terminal portion. In hair samples taken 5 years after death, no significant changes in morphology were observed. By 15 years after death, however, we observed losses in various layers of the hair, including the cuticle layer. At 25 years after death, hair shafts showed a number of pores extending into the medulla, with only some hair shafts retaining their cortical layers. To our knowledge, this is the first ultramicroscopic study on weathering of hair for up to 25 years after death. Our results may therefore provide a basis for similar studies in the fields of forensic science and physical anthropology.