Molecular Dynamics Investigation into CerENP's Effect on the Lipid Matrix of Stratum Corneum.

The extracellular lipid matrix in the stratum corneum (SC) plays a critical role in skin barrier functionality, comprising three primary components: ceramides, cholesterol, and free fatty acids. The diverse ceramides, differentiated by molecular structures such as hydroxylations and varying chain lengths, are essential for the lipid matrix's structural integrity. Recently, a new subclass of ceramide, 1-O-acylceramide NP (CerENP), has been identified; however, its precise role in the lipid matrix of the SC is still elusive. Herein, we investigate the role of CerENP on the structure and permeability of the SC using molecular dynamics simulations. Our findings indicate that CerENP contributes to a compact lipid matrix in the lateral dimension of our SC model with a repeat distance of about 13 nm. Additionally, ethanol permeability assessments show that CerENP effectively reduces molecular penetration through the lipid matrix. This study provides an insight into the role of a new subclass of ceramide in the SC, enhancing our understanding of skin structure and the mechanisms behind barrier dysfunction in skin diseases.

Anisotropically Conductive Hydrogels with Directionally Aligned PEDOT:PSS in a PVA Matrix.

Electrical anisotropy, which is characterized by the efficient transmission of electrical signals in specific directions, is prevalent in both natural and engineered systems. However, traditional anisotropically conductive materials are often rigid and dry, thus limiting their utility in applications aiming for the seamless integration of various technologies with biological tissues. In the present study, we introduce a method for precisely controlling the microstructures of conductive and insulating polymers to create highly anisotropically conductive composite hydrogels. Our methodology involves combining aligned poly(vinyl alcohol) microfibrils, infused poly(3,4-ethylenedioxythiophene) polystyrenesulfonate, and sodium citrate precipitation to form dense, aligned conductive paths. This significantly enhances the electrical conductivity anisotropy (σ∥/σ⊥ ≈ 60.8) within these composite hydrogels.

Skin hydration modulatory activities of Ficus deltoidea extract.

Ficus deltoidea was known for its potent antioxidant, anti-melanogenic and photoprotective skin barrier activities. These properties are contributed by its biomarkers which are vitexin and isovitexin. This study aims to optimize the yield of methanolic extraction of Ficus deltoidea leaves (EFD) and evaluate their effects on skin barrier function and hydration. For optimization, Box-Behnken design was utilized to investigate the effects of methanol concentration, sonication time, and solvent-to-sample ratio on the yields of vitexin and isovitexin in EFD. The optimal yields obtained were 32.29 mg/g for vitexin and 35.87 mg/g for isovitexin. The optimum extraction conditions were 77.66% methanol concentration, 20.03 min sonication time, and 19.88 mL/g solvent-to-sample ratio. The quantitative real-time polymerase chain reaction was utilized to measure variant marker genes of transglutaminase-1, caspase 14, ceramide synthase 3, involucrin, and filaggrin of EFD-induced keratinocyte differentiation by in vitro study. Exposure to EFD has elevated the mRNA levels of all tested marker genes by 0.7-9.2 folds. Then, in vivo efficacy study was conducted on 20 female subjects for 14 days to evaluate skin biophysical assessment of hydration. EFD topical formulation treatment successfully increased skin hydration on day 7 (43.74%) and day 14 (47.23%). In silico study by molecular docking was performed to identify intermolecular binding interactions of vitexin and isovitexin with the interested proteins of tested marker genes. The result of molecular docking to the interested proteins revealed a similar trend with real-time PCR data. In conclusion, EFD potentially enhanced the skin barrier function and hydration of human skin cells.

Hydrogel-based strong and fast actuators by electroosmotic turgor pressure.

Hydrogels are promising as materials for soft actuators because of qualities such as softness, transparency, and responsiveness to stimuli. However, weak and slow actuations remain challenging as a result of low modulus and osmosis-driven slow water diffusion, respectively. We used turgor pressure and electroosmosis to realize a strong and fast hydrogel-based actuator. A turgor actuator fabricated with a gel confined by a selectively permeable membrane can retain a high osmotic pressure that drives gel swelling; thus, our actuator exerts large stress [0.73 megapascals (MPa) in 96 minutes (min)] with a 1.16 cubic centimeters of hydrogel. With the accelerated water transport caused by electroosmosis, the gel swells rapidly, enhancing the actuation speed (0.79 MPa in 9 min). Our strategies enable a soft hydrogel to break a brick and construct underwater structures within a few minutes.

Ceramide NPs derived from natural oils of Korean traditional plants enhance skin barrier functions and stimulate expressions of genes for epidermal homeostasis.

BACKGROUND: New ceramide (CER) NPs were prepared by linking fatty acids derived from oils of Korean traditional plants to phytosphingosine (PHS). The oils of Korean traditional plants were extracted from the seeds of Panax ginseng, Camellia sinensis, Glycine max napjakong, Glycine max seoritae, and Camellia japonica as sources of diverse fatty acids. AIMS: The aim of this study was to investigate signaling bioactivities of HP-C. sinensis ceramide NP that was column purified to remove any residual PHS and to evaluate the skin barrier functions of the HP-C. sinensis ceramide NP in human skin. METHODS: The expressions of genes related to epidermal differentiation were analyzed in vitro by qPCR. Human studies were also performed to determine the skin barrier functions with respect to TEWL and SC cohesion. RESULTS: The HP-C. sinensis CER NP significantly enhanced the expressions of FLG, CASP14, and INV indicates that the signaling biological activities of oil-derived ceramide NPs could be different depend on the natural oils. The control ceramide, C18-CER NP, had no effect on the expression of the three genes. HP-C. sinensis CER NP was selected for the in vivo human studies. Application of 0.5% HP-C. sinensis CER NP cream stimulated significantly faster recovery of a disrupted skin barrier than that of the control C18-CER NP. A significant enhancement of SC cohesion of the skin treated with 0.5% HP-C. sinensis CER NP was also observed. CONCLUSION: Taken all together, our results clearly demonstrate that HP-C. sinensis CER NP, P. ginseng CER NP, and other oil-derived CER NP could be a better choice for developing moisturizers to improve skin barrier function as they more closely mimic the endogenous CER composition of the actual human skin barrier.

Identification of peptide inhibitors of matrix metalloproteinase 1 using an in-house assay system for the enzyme.

Matrix metalloproteinases (MMPs) are zinc-dependent proteases involved in the degradation of extracellular matrix proteins. As one of the isoforms, MMP-1 breaks down collagen, and its activity is known to be important in wound healing. Its timely and adequate level of expression is pivotal because MMP-1 is also involved in the damage or aging of skins as well as in certain types of cancers. Thus, both assaying the MMP-1 activity and developing its inhibitors are of great importance. We here developed an in-house assay system that gave us the high degree of freedom in screening peptide inhibitors of MMP-1. The assay system utilized a circularly permutated fusion of ß-lactamase and its inhibitory protein through an MMP-1-sensitive linker so that the activity of MMP-1 could be translated into that of ß-lactamase. As a proof of concept, we applied the developed assay system to initial screens of MMP-1 inhibitors and successfully identified one lead peptide that inhibited the collagenase activity of the enzyme.

Phytosphingosine Increases Biosynthesis of Phytoceramide by Uniquely Stimulating the Expression of Dihydroceramide C4-desaturase (DES2) in Cultured Human Keratinocytes.

Ceramide NP is known to be the most abundant class of 12 ceramide (CER) families that form a permeability barrier in the human skin barrier. However, not many studies have been reported on the regulation of the biosynthesis of ceramide NP. Recently, it has been reported that phytosphingosine (PHS) treatment in the cultured keratinocytes (KC) notably increased the content of ceramide NP. However, the mechanism behind the PHS-induced enhancement of ceramide NP has not been elucidated. In this study, we investigated the effects of PHS on the expression of several essential genes for the biosynthesis of CER. Also, we determined the molecular mechanism behind the unique enhancement of ceramide NP upon treatment of PHS in the cultured KC. The expressions of all of the three genes (SPT, ceramide synthase 3 [CERS3], and ELOVL4) and their respective proteins were markedly increased in PHS-treated KC. In addition, the expression of the dihydroceramide C4-desaturase (DES2) responsible for conversion of dihydroceramide into ceramide NP was uniquely enhanced only by PHS treatment. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis revealed that more than 20-fold increase of ceramide NP by PHS was observed while no significant enhancement of ceramide NS and NDS was observed. This study demonstrates that PHS plays a fundamental role in strengthening the epidermal permeability barrier by stimulating the overall processes of biosynthesis of all classes of CER in epidermis. The dramatic increase of ceramide NP upon PHS treatment seemed to be the outcome of transformation of dihydroceramide and/or ceramide NS by C4-hydroxylase activity.

Impaired permeability and antimicrobial barriers in type 2 diabetes skin are linked to increased serum levels of advanced glycation end-product.

The incidence of type 2 diabetes mellitus (DM) has been increasing rapidly, and the disease has become a serious sociomedical problem. Many skin problems, such as xerosis, pruritus, skin infections and delayed wound healing, that might be related to chronic impairment of skin barrier function decrease the quality of life in patients with DM. However, the status of the permeability and antimicrobial barrier of the skin in DM remains unknown. This study aimed to elucidate skin barrier impairment in patients with type 2 DM and its pathomechanisms using classic animal models of type 2 DM. Functional studies of the skin barrier and an analysis of stratum corneum (SC) lipids were compared between patients with type 2 DM and age- and sex-matched non-diabetes controls. Also, functional studies on the skin barrier, epidermal lipid analyses, and electron microscopy and biomolecular studies were performed using type 2 DM animal models, db/db and ob/ob mice. Patients with type 2 DM presented with epidermal barrier impairments, including SC hydration, which was influenced by blood glucose control (HbA1c level). In the lipid analysis of SC, ceramides, fatty acids and cholesterol were significantly decreased in patients with type 2 DM compared with controls. Type 2 DM murine models presented with severe hyperglycaemia, impairment of skin barrier homeostasis, decreases in epidermal proliferation and epidermal lipid synthesis, decreases in lamellar body (LB) and epidermal antimicrobial peptides (AMPs), an increase in receptors for advanced glycation end-product (AGE) in the epidermis and an increase in serum AGE. Impairment of the skin barrier was observed in type 2 DM, which results in part from a decrease in epidermal proliferation. Serum AGE and its epidermal receptors were increased in type 2 diabetic mice which display impaired skin barrier parameters such as epidermal lipid synthesis, LB production, epidermal AMP and SC lipids.

Novel phytoceramides containing fatty acids of diverse chain lengths are better than a single C18-ceramide N-stearoyl phytosphingosine to improve the physiological properties of human stratum corneum.

Ceramides in the human stratum corneum (SC) are a mixture of diverse N-acylated fatty acids (FAs) with different chain lengths. C24 is the major class of FAs of ceramides. However, there are also other classes of ceramides with diverse chain lengths of FAs, and these lengths generally range from C16 to C26. This study aimed to prepare several types of phytoceramide containing diverse chain lengths of N-acylated FAs and compare them with C18-ceramide N-stearoyl phytosphingosine (NP) in terms of their effects on the physiological properties of the SC. We chose natural oils, such as horse fat oil, shea butter, sunflower oil, and a mixture of macadamia nut, shea butter, moringa, and meadowfoam seed oil, as sources of FAs and phytosphingosine as a sphingoid backbone to synthesize diverse phytoceramides. Each phytoceramide exhibited a distinctive formation of the lamellar structure, and their FA profiles were similar to those of their respective natural oil. The skin barrier properties, as analyzed in human skin, clearly demonstrated that all the phytoceramides improved the recovery rate of the damaged SC and enhanced hydration better than C18-ceramide NP did. In conclusion, natural oil-derived phytoceramides could represent a novel class of ceramides for cosmetic applications in the development of an ideal skin barrier moisturizer.

Phytosphingosine enhances moisture level in human skin barrier through stimulation of the filaggrin biosynthesis and degradation leading to NMF formation.

Phytosphingosine (PHS) is a sphingoid that is a key component of phytoceramides NP, AP and EOP. PHS has been known to have anti-inflammation and antimicrobial activities and to stimulate epidermal differentiation. In addition, it is reported that PHS treatment notably increased phytoceramide content in keratinocytes. In this study, we tried to investigate whether PHS has any effect on the maturation of corneocytes such as formation of cornified envelope and natural moisturizing factor (NMF) that is also an essential event during the formation of skin barrier, stratum corneum. Special focus was made on the filaggrin (FLG) metabolism that is directly responsible for NMF production. PHS increased the expression of essential keratinocyte differentiation genes such as involucrin and transglutaminase 1 in cultured human keratinocytes. Interestingly, the expressions of FLG, caspase 14 and bleomycin hydrolase, all of which involved in NMF production in corneocytes, were significantly induced by PHS treatment in vitro. The effect of PHS on FLG metabolism was manifested as the increase of pyrrolidone carboxylic acid and skin hydration in vivo human skin. Results showed PHS had skin moisturizing effect by modulating FLG metabolic pathways and suggested to be an essential role in coordinated formation of the corneocyte envelope and NMF within.

Reduced Nrf2 activation in PI3K phosphorylation-impaired vitiliginous keratinocytes increases susceptibility to ROS-generating chemical-induced apoptosis.

Keratinocytes in affected epidermis of vitiligo patients are known to have impaired activation of the PI3K/AKT pathway. Based on critical roles of keratinocytes and oxidative stress in vitiligo development, this study examined whether keratinocytes with impaired PI3K activation were more vulnerable to apoptosis caused by oxidative stress from phenolic compounds, p-tert-butylphenol (4-TBP) and hydroquinone (HQ). Cell viability assay, FACS analysis, ELISA for TNF-α or IL-1a, ROS assay, Western blot analysis for Nrf2 expression, and confocal microscopy with anti-Nrf2 and phospho-PI3K antibodies were done on primary cultured normal human keratinocytes with or without PI3K knockdown in the presence or absence of chemical treatment or antioxidant. Immunofluorescence staining using anti-Nrf2, phospho-PI3K, TNF-ɑ, and IL-1ɑ antibodies, ROS assay using dihydroethidium, and TUNEL assay were performed on sets of depigmented and normally pigmented skin from vitiligo patients. Results showed that 4-TBP or HQ treatment increased apoptosis and the expression levels of TNF-ɑ, IL-1ɑ, and ROS in PI3K-knockdown keratinocytes which reduced Nrf2 nuclear translocation compared to control keratinocytes. These changes were significantly recovered by an antioxidant treatment. Depigmented epidermis of vitiligo patients also showed lower levels of Nrf2 and phospho-PI3K but higher levels of ROS, TNF-ɑ, IL-1ɑ, and ROS with more TUNEL-positive cells. Therefore, impaired PI3K activation in keratinocytes in depigmented epidermis of vitiligo patients are vulnerable to apoptosis caused by ROS-generating chemicals due to reduced Nrf2 activation.

Inhibitory Effects of Novel SphK2 Inhibitors on Migration of Cancer Cells.

BACKGROUND: Cell migration is an essential process for survival and differentiation of mammalian cells. Numerous diseases are induced or influenced by inappropriate regulation of cell migration, which plays a key role in cancer cell metastasis. In fact, very few anti-metastasis drugs are available on the market. SphKs are enzymes that convert sphingosine to sphingosine-1-phosphate (S1P) and are known to control various cellular functions, including migration of cells. In human, SphK2 is known to promote apoptosis, suppresses cell growth, and controls cell migration; in addition, the specific ablation of SphK2 activity was reported to inhibit cancer cell metastasis. OBJECTIVE: The previously identified SG12 and SG14 are synthetic analogs of sphingoid and can specifically inhibit the functions of SphK2. We investigated the effects of the SphK2 specific inhibitors on the migratory behavior of cells. METHOD: We investigated how SG12 and SG14 affect cell migration by monitoring both cumulative and individual cell migration behavior using HeLa cells. RESULTS: SG12 and SG14 mutually showed stronger inhibitory effects with less cytotoxicity compared with a general SphK inhibitor, N,N-dimethylsphingosine (DMS). The mechanistic aspects of specific SphK2 inhibition were studied by examining actin filamentation and the expression levels of motility-related genes. CONCLUSION: The data revealed that SG12 and SG14 resemble DMS in decreasing overall cell motility, but differ in that they differentially affect motility parameters and motility-related signal transduction pathways and therefore actin polymerization, which are not altered by DMS. Our findings show that SphK2 inhibitors are putative candidates for anti-metastatic drugs.

A synthetic C16 omega-hydroxyphytoceramide improves skin barrier functions from diversely perturbed epidermal conditions.

Omega-hydroxyceramides (ω-OH-Cer) play a crucial role in maintaining the integrity of skin barrier. ω-OH-Cer are the primary lipid constituents of the corneocyte lipid envelope (CLE) covalently attached to the outer surface of the cornified envelope linked to involucrin to become bound form lipids in stratum corneum (SC). CLE becomes a hydrophobic impermeable layer of matured corneocyte preventing loss of natural moisturizing factor inside the corneocytes. More importantly, CLE may also play an important role in the formation of proper orientation of intercellular lipid lamellar structure by interdigitating with the intercellular lipids in a comb-like fashion. Abnormal barrier conditions associated with atopic dermatitis but also UVB-irradiated skins are known to have lowered level of bound lipids, especially ω-OH-Cer, which indicate that ω-OH-Cer play an important role in maintaining the integrity of skin barrier. In this study, protective effects of a novel synthetic C16 omega-hydroxyphytoceramides (ω-OH-phytoceramide) on skin barrier function were investigated. Epidermal barrier disruption was induced by UVB irradiation, tape-stripping in hairless mouse and human skin. Protective effect of damaged epidermis was evaluated using the measurement of transepidermal water loss and cohesion of SC. Increased keratinocyte differentiation was verified using cultured keratinocyte through western blot. Results clearly demonstrated that a synthetic C16 ω-OH-phytoceramide enhanced the integrity of SC and accelerated the recovery of damaged skin barrier function by stimulating differentiation process. In a conclusion, a synthetic C16 ω-OH-phytoceramide treatment improved epidermal homeostasis in several disrupted conditions.

Lipidomic platform for structural identification of skin ceramides with α-hydroxyacyl chains.

Skin ceramides are sphingolipids consisting of sphingoid bases, which are linked to fatty acids via an amide bond. Typical fatty acid acyl chains are composed of α-hydroxy fatty acid (A), esterified ω-hydroxy fatty acid (EO), non-hydroxy fatty acid (N), and ω-hydroxy fatty acid (O). We recently established a lipidomic platform to identify skin ceramides with non-hydroxyacyl chains using tandem mass spectrometry. We expanded our study to establish a lipidomic platform to identify skin ceramides with α-hydroxyacyl chains. Tandem mass spectrometry analysis of A-type ceramides using chip-based direct infusion nanoelectrospray-mass spectrometry showed the characteristic fragmentation pattern of both acyl and sphingoid units, which can be applied for structural identification of ceramides. Based on the tandem mass spectrometry fragmentation patterns of A-type ceramides, comprehensive fragmentation schemes were proposed. Our results may be useful for identifying A-type ceramides in the stratum corneum of human skin.

A nuclear factor kappa B-derived inhibitor tripeptide inhibits UVB-induced photoaging process.

BACKGROUND: Ultraviolet (UV) irradiation on the skin induces photoaging which is characterized by keratinocyte hyperproliferation, generation of coarse wrinkles, worse of laxity and roughness. Upon UV irradiation, nuclear factor kappa B (NF-κB) is activated which plays a key role in signaling pathway leading to inflammation cascade and this activation stimulates expression of pro-inflammatory cytokines such as tumor necrosis factor alpha (TNF-α), interleukin-1alpha (IL-1α) and a stress response gene cyclooxygenase-2 (COX-2). In addition, activation of NF-κB up-regulates the expression of matrix metalloprotease-1 (MMP-1) and consequently collagen in dermis is degraded. OBJECTIVE: In this study, the effects of a NF-κB-derived inhibitor tripeptide on the UVB-induced photoaging and inflammation were investigated in vitro and in vivo. METHODS: A NF-κB-derived inhibitor tripeptide (NF-κB-DVH) was synthesized based on the sequence of dimerization region of the subunit p65 of NF-κB. Its inhibitory activity was confirmed using chromatin immunoprecipitation assay and in situ proximate ligation assay. The effects of anti-photoaging and anti-inflammation were analyzed by Enzyme-linked immunosorbent assay (ELISA), immunoblotting and immunochemistry. RESULTS: NF-κB-DVH significantly decreased UV-induced expression of TNF-α, IL-1α, MMP-1 and COX-2 while increased production of type I procollagen. CONCLUSION: Results showed NF-κB-DVH had strong anti-inflammatory activity probably by inhibiting NF-κB activation pathway and suggested to be used as a novel agent for anti-photoaging.

Factors influencing superimposition error of 3D cephalometric landmarks by plane orientation method using 4 reference points: 4 point superimposition error regression model.

Superimposition has been used as a method to evaluate the changes of orthodontic or orthopedic treatment in the dental field. With the introduction of cone beam CT (CBCT), evaluating 3 dimensional changes after treatment became possible by superimposition. 4 point plane orientation is one of the simplest ways to achieve superimposition of 3 dimensional images. To find factors influencing superimposition error of cephalometric landmarks by 4 point plane orientation method and to evaluate the reproducibility of cephalometric landmarks for analyzing superimposition error, 20 patients were analyzed who had normal skeletal and occlusal relationship and took CBCT for diagnosis of temporomandibular disorder. The nasion, sella turcica, basion and midpoint between the left and the right most posterior point of the lesser wing of sphenoidal bone were used to define a three-dimensional (3D) anatomical reference co-ordinate system. Another 15 reference cephalometric points were also determined three times in the same image. Reorientation error of each landmark could be explained substantially (23%) by linear regression model, which consists of 3 factors describing position of each landmark towards reference axes and locating error. 4 point plane orientation system may produce an amount of reorientation error that may vary according to the perpendicular distance between the landmark and the x-axis; the reorientation error also increases as the locating error and shift of reference axes viewed from each landmark increases. Therefore, in order to reduce the reorientation error, accuracy of all landmarks including the reference points is important. Construction of the regression model using reference points of greater precision is required for the clinical application of this model.

A lipidomic platform establishment for structural identification of skin ceramides with non-hydroxyacyl chains.

The stratum corneum (SC) is the outermost layer of skin that functions as a barrier and protects against environmental influences and transepidermal water loss. Its unique morphology consists of keratin-enriched corneocytes embedded in a distinctive mixture of lipids containing mainly ceramides, free fatty acids, and cholesterol. Ceramides are sphingolipids consisting of sphingoid bases, which are linked to fatty acids by an amide bond. Typical sphingoid bases in the skin are composed of dihydrosphingosine (dS), sphingosine (S), phytosphingosine (P), and 6-hydroxysphingosine (H), and the fatty acid acyl chains are composed of non-hydroxy fatty acid (N), α-hydroxy fatty acid (A), ω-hydroxy fatty acid (O), and esterified ω-hydroxy fatty acid (E). The 16 ceramide classes include several combinations of sphingoid bases and fatty acid acyl chains. Among them, N-type ceramides are the most abundant in the SC. Mass spectrometry (MS)/MS analysis of N-type ceramides using chip-based direct infusion nanoelectrospray-ion trap mass spectrometry generated the characteristic fragmentation pattern of both acyl and sphingoid units, which could be applied to structural identification of ceramides. Based on the MS/MS fragmentation patterns of N-type ceramides, comprehensive fragmentation schemes were proposed. In addition, mass fragmentation patterns, which are specific to the sphingoid backbone of N-type ceramides, were found in higher m/z regions of tandem mass spectra. These characteristic and general fragmentation patterns were used to identify N-type ceramides in human SC. Based on established MS/MS fragmentation patterns of N-type ceramides, 52 ceramides (including different classes of NS, NdS, NP, and NH) were identified in human SC. The MS/MS fragmentation patterns of N-type ceramides were characterized by interpreting their product ion scan mass spectra. This information may be used to identify N-type ceramides in the SC of human, rat, and mouse skin.

Cone beam CT findings of retromolar canals: Report of cases and literature review.

A retromolar canal is an anatomical variation in the mandible. As it includes the neurovascular bundle, local anesthetic insufficiency can occur, and an injury of the retromolar canal during dental surgery in the mandible may result in excessive bleeding, paresthesia, and traumatic neuroma. Using imaging analysis software, we evaluated the cone-beam computed tomography (CT) images of two Korean patients who presented with retromolar canals. Retromolar canals were detectable on the sagittal and cross-sectional images of cone-beam CT, but not on the panoramic radiographs of the patients. Therefore, the clinician should pay particular attention to the identification of retromolar canals by preoperative radiographic examination, and additional cone beam CT scanning would be recommended.

Comparison of conventional lateral cephalograms with corresponding CBCT radiographs.

PURPOSE: This study was performed to assess the compatibility of cone beam computed tomography (CBCT) synthesized cephalograms with conventional cephalograms, and to find a method for obtaining normative values for three-dimensional (3D) assessments. MATERIALS AND METHODS: The sample group consisted of 10 adults with normal occlusion and well-balanced faces. They were imaged using conventional and CBCT cephalograms. The CBCT cephalograms were synthesized from the CBCT data using OnDemand 3D software. Twenty-one angular and 12 linear measurements from each imaging modality were compared and analyzed using paired-t test. RESULTS: The linear measurements between the two imaging modalities were not statistically different (p>0.05) except for the U1 to facial plane distance. The angular measurements between the two imaging modalities were not statistically different (p>0.05) with the exception of the gonial angle, ANB difference, and facial convexity. CONCLUSION: Two-dimensional cephalometric norms could be readily used for 3D quantitative assessment, if corrected for lateral cephalogram distortion.