Functional characterization of O-methyltransferases used to catalyse site-specific methylation in the post-tailoring steps of pradimicin biosynthesis.

AIMS: To identify the roles of the two O-methyltransferase homologous genes pdmF and pdmT in the pradimicin biosynthetic gene cluster of Actinomadura hibisca P157-2. METHODS AND RESULTS: Pradimicins are pentangular polyphenol antibiotics synthesized by bacterial type II polyketide synthases (PKSs) and tailoring enzymes. Pradimicins are naturally derivatized by combinatorial O-methylation at two positions (i.e., 7-OH and 11-OH) of the benzo[α]naphthacenequinone structure. PdmF and PdmT null mutants (PFKO and PTKO) were generated. PFKO produced the 11-O-demethyl shunt metabolites 11-O-demethylpradimicinone II (1), 11-O-demethyl-7-methoxypradimicinone II (2), 11-O-demethylpradimicinone I (3) and 11-O-demethylpradimicin A (4), while PTKO generated the 7-O-demethyl derivatives pradimicinone II (5) and 7-hydroxypradimicin A (6). Pradimicinones 1, 2, 3, and 5 were fed to a heterologous host Escherichia coli harbouring expression plasmid pET-22b::pdmF or pET-28a::pdmT. PdmF catalysed 11-O-methylation of pradimicinones 1, 2, and 3 regardless of O-methylation at the C-7 position, while PdmT was unable to catalyse 7-O-methylation when the C-11 hydroxyl group was methylated (5). CONCLUSIONS: PdmF and PdmT were involved in 11-O- and 7-O-methylations of the benzo[α]naphthacenequinone moiety of pradimicin, respectively. Methylation of the C-7 hydroxyl group precedes methylation of the C-11 hydroxyl group in pradimicin biosynthesis. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first reported demonstration of the functions of PdmF and PdmT for regiospecific O-methylation, which contributes to better understanding of the post-PKS modifications in pradimicin biosynthesis as well as to rational engineering of the pradimicin biosynthetic machinery.

Association of lipid parameters and insulin resistance with bone health in South Korean adolescents.

UNLABELLED: This study investigated the association between lipid profiles and insulin resistance and bone mineral content (BMC) in Korean adolescents and found that BMC was inversely associated with triglyceride (TG) and homeostasis model assessment of insulin resistance (HOMA-IR). This association did not differ according to obesity status in either boys or girls. INTRODUCTION: To prevent future osteoporosis, it is important to identify factors that affect bone health in adolescents as well as adults. This study aimed to examine the association between lipid profiles and insulin resistance and BMC in Korean adolescents. METHODS: Data from 706 boys and 621 girls, who participated in the Korea National Health and Nutrition Examination Survey from 2008 to 2011, were analyzed. Lipid profiles were measured, and HOMA-IR was calculated to assess insulin resistance. BMC was measured for the total femur, femur neck, and lumbar spine by using whole-body dual-energy X-ray absorptiometry (DXA). RESULTS: TG level and HOMA-IR were negatively correlated with BMC at all three sites in boys. In girls, TG level showed a negative correlation with BMC at the femur neck and lumbar spine, and HOMA-IR was negatively associated with BMC at the femur neck only. These inverse associations did not differ according to obesity status in either sex. Adjusted means of BMC at the three sites in boys tended to decrease in the higher tertile groups of TG and HOMA-IR, and the adjusted means of BMC for the total femur in girls tended to decrease in the higher tertile groups of TG and HOMA-IR. CONCLUSIONS: BMC was inversely associated with TG and HOMA-IR in Korean adolescents, and this association was more pronounced in boys. This association did not differ according to obesity status in either sex.

Regio-selectively reduced streptogramin A analogue, 5,6-dihydrovirginiamycin M1 exhibits improved potency against MRSA.

A newly reduced macrocyclic lactone antibiotic streptogramin A, 5,6-dihydrovirginiamycin M1 was created by feeding virginiamycin M1 into a culture of recombinant Streptomyces venezuelae. Its chemical structure was spectroscopically elucidated, and this streptogramin A analogue showed twofold higher antibacterial activities against methicillin-resistant Staphylococcus aureus (MRSA) compared with its parent molecule virginiamycin M1. Docking studies using the model of streptogramin A acetyltransferase (VatA) suggested that the newly generated analogue binds tighter with overall lower free energy compared with the parent molecule virginiamycin M1. This hypothesis was validated experimentally through the improvement of efficacy of the new analogue against MRSA strains. The biotransformation approach presented herein could have a broad application in the production of reduced macrocyclic molecules.

Retroductal Delivery of Epidermal Growth Factor Protects Salivary Progenitors after Irradiation.

Salivary gland hypofunction after irradiation is associated with a deficit of epithelial stem/progenitors in salivary glands. Although epidermal growth factor (EGF) is known to stimulate the proliferation of epithelial cells, the therapeutic effect of EGF on salivary epithelial stem/progenitors remains undetermined. In this study, we administered EGF to submandibular glands (SMGs) via a retrograde route through the SMG excretory duct before fractionated irradiation and examined whether EGF could protect salivary epithelial progenitor cells from radiation and alleviate radiation-induced salivary hypofunction. EGF-treated mice exhibited greater body and gland weights at 12 wk after irradiation than untreated mice. The retroductal delivery of EGF improved salivary secretory function and increased salivary amylase activity in a dose-dependent manner. Histological examinations highlighted the amelioration of the loss of keratine-14+ (KRT14+) basal ductal and/or MIST1+ acinar cells, as well as induction of fibrosis, following irradiation in EGF-treated mice. An additional in vitro experiment using a salivary gland organoid irradiation model indicated that the radioprotective effects of EGF promoted the growth and inhibited the apoptotic cell death of salivary epithelial cells. Our results suggest that retroductal delivery of EGF may be a promising therapeutic option for preventing radiation-induced salivary gland hypofunction.

Performance of electron beam irradiation for treatment of groundwater contaminated with acetone.

The purposes of this study were to evaluate the efficiency of acetone removal by electron beam irradiation in groundwater and the effect of various conditions. According to the results, the removal kinetics of acetone were pseudo first-order, and the removal efficiencies were expressed to the (%) removal and G-values. By adding sulfite, it was confirmed that acetone was mainly degraded by the reaction with the hydrated electrons. The presence of nitrate caused the removal of acetone to decrease. But there was no significant effect of alkalinity on the removal of acetone. The effect of the initial pH values (pH 5 to 9) on the acetone removal efficiency was negligible, but the pH value decreases due to the formation of acidic compounds after irradiation. Consequently, the radiation-induced removal reactions of acetone followed the pseudo-first-order kinetic model; in addition to the initial concentration of acetone, nitrate and the absorbed dose were important factors in removing acetone from an aqueous solution using electron beam irradiation. The effects of general pH and alkalinity on the degrading acetone were negligible.

In vitro metabolism of a novel PPAR gamma agonist, KR-62980, and its stereoisomer, KR-63198, in human liver microsomes and by recombinant cytochrome P450s.

1. KR-62980 and its stereoisomer KR-63198 are novel and selective peroxisome proliferator-activated receptor gamma (PPAR gamma) modulators with activity profiles different from that of rosiglitazone. This study was performed to identify the major metabolic pathways for KR-62980 and KR-63198 in human liver microsomes. 2. Human liver microsomal incubation of KR-62980 and KR-63198 in the presence of a beta-nicotinamide adenine dinucleotide phosphate (NADPH)-generating system resulted in hydroxy metabolite formation. In addition, the specific cytochrome P450s (CYPs) responsible for KR-62980 and KR-63198 hydroxylation were identified by using a combination of chemical inhibition in human liver microsomes and metabolism by recombinant P450s. It is shown that CYP1A2, CYP2D6, CYP3A4, and CYP3A5 are the predominant enzymes in the hydroxylation of KR-62980 and KR-63198. 3. The intrinsic clearance through hydroxylation was consistently and significantly higher for KR-62980 than for KR-63198, indicating metabolic stereoselectivity (CL(int) of 0.012 +/- 0.001 versus 0.004 +/- 0.001 microl min(-1) pmol(-1) P450, respectively). 4. In a drug-drug interaction study, KR-62980 and KR-63198 had no effect on the activities of the P450s tested (IC(50) > 50 microM), suggesting that in clinical interactions between KR-62980 and KR-63198 the P450s tested would not be expected.

RNA polymerase beta-subunit gene (rpoB) sequence analysis for the identification of Bacteroides spp.

Partial rpoB sequences (317 bp) of 11 species of Bacteroides, two Porphyromonas spp. and two Prevotella spp. were compared to delineate the genetic relationships among Bacteroides and closely related anaerobic species. The high level of inter-species sequence dissimilarities (7.6-20.8%) allowed the various Bacteroides spp. to be distinguished. The position of the Bacteroides distasonis and Bacteriodes merdae cluster in the rpoB tree was different from the position in the 16S rRNA gene tree. Based on rpoB sequence similarity and clustering in the rpoB tree, it was possible to correctly re-identify 80 clinical isolates of Bacteroides. In addition to two subgroups, cfiA-negative (division I) and cfiA-positive (division II), of Bacteroides fragilis isolates, two distinct subgroups were also found among Bacteroides ovatus and Bacteroides thetaiotaomicron isolates. Bacteroides genus-specific rpoB PCR and B. fragilis species-specific rpoB PCR allowed Bacteroides spp. to be differentiated from Porphyromonas and Prevotella spp., and also allowed B. fragilis to be differentiated from other non-fragilisBacteroides spp. included in the present study.

A novel scratching approach for measuring age-related changes in the in situ toughness of bone.

A scratch test using a nanoindentation system was proposed in this study to assess the age-related changes in the in situ toughness of bone matrix at ultrastructural levels. A tissue removal energy density (u(r)) was defined and estimated as the work done by the scratch (U(T)) divided by the total volume of the scratch groove (u(s)). The value of u(s) was used as a relative measure of the in situ toughness of the tissue. Human cortical bone specimens obtained from middle-aged (between 49 and 59 years old) and elderly groups (over 69 years old) were tested using this technique. A significant difference in the estimated removal energy density (u(s)) in the secondary osteons was found between the middle-aged and elderly groups (5.49+/-0.696 vs. 4.09+/-1.30 N/mm(2), respectively).

Arctic sea ice melt leads to atmospheric new particle formation.

Atmospheric new particle formation (NPF) and growth significantly influences climate by supplying new seeds for cloud condensation and brightness. Currently, there is a lack of understanding of whether and how marine biota emissions affect aerosol-cloud-climate interactions in the Arctic. Here, the aerosol population was categorised via cluster analysis of aerosol size distributions taken at Mt Zeppelin (Svalbard) during a 11 year record. The daily temporal occurrence of NPF events likely caused by nucleation in the polar marine boundary layer was quantified annually as 18%, with a peak of 51% during summer months. Air mass trajectory analysis and atmospheric nitrogen and sulphur tracers link these frequent nucleation events to biogenic precursors released by open water and melting sea ice regions. The occurrence of such events across a full decade was anti-correlated with sea ice extent. New particles originating from open water and open pack ice increased the cloud condensation nuclei concentration background by at least ca. 20%, supporting a marine biosphere-climate link through sea ice melt and low altitude clouds that may have contributed to accelerate Arctic warming. Our results prompt a better representation of biogenic aerosol sources in Arctic climate models.

Biosynthesis of the ansamycin antibiotic rifamycin: deductions from the molecular analysis of the rif biosynthetic gene cluster of Amycolatopsis mediterranei S699.

BACKGROUND: The ansamycin class of antibiotics are produced by various Actinomycetes. Their carbon framework arises from the polyketide pathway via a polyketide synthase (PKS) that uses an unusual starter unit. Rifamycin (rif), produced by Amycolatopsis mediterranei, is the archetype ansamycin and it is medically important. Although its basic precursors (3-amino-5-hydroxy benzoic acid AHBA, and acetic and propionic acids) had been established, and several biosynthetic intermediates had been identified, very little was known about the origin of AHBA nor had the PKS and the various genes and enzymes that modify the initial intermediate been characterized. RESULTS: A set of 34 genes clustered around the rifK gene encoding AHBA synthase were defined by sequencing all but 5 kilobases (kb) of a 95 kb contiguous region of DNA from A. mediterranei. The involvement of some of the genes in the biosynthesis of rifamycin B was examined. At least five genes were shown to be essential for the synthesis of AHBA, five genes were determined to encode the modular type I PKS that uses AHBA as the starter unit, and 20 or more genes appear to govern modification of the polyketide-derived framework, and rifamycin resistance and export. Putative regulatory genes were also identified. Disruption of the PKS genes at the end of rifA abolished rifamycin B production and resulted in the formation of P8/1-OG, a known shunt product of rifamycin biosynthesis, whereas disruption of the orf6 and orf9 genes, which may encode deoxysugar biosynthesis enzymes, had no apparent effect. CONCLUSIONS: Rifamycin production in A. mediterranei is governed by a single gene cluster consisting of structural, resistance and export, and regulatory genes. The genes characterized here could be modified to produce novel forms of the rifamycins that may be effective against rifamycin-resistant microorganisms.

Characterization of the enzymatic domains in the modular polyketide synthase involved in rifamycin B biosynthesis by Amycolatopsis mediterranei.

Five clustered polyketide synthase (PKS) genes, rifA-rifE, involved in rifamycin (Rf) biosynthesis in Amycolatopsis mediterranei S699 have been cloned and sequenced (August, P.R. et al., 1998. Chem. Biol. 5, 69-79). The five multifunctional polypeptides constitute a type I modular PKS that contains ten modules, each responsible for a specific round of polyketide chain elongation. Sequence comparisons of the Rf PKS proteins with other prokaryotic modular PKSs elucidated the regions that have an important role in enzyme activity and specificity. The beta-ketoacyl:acyl carrier protein synthase (KS) domains show the highest degree of similarity between themselves (86-90%) and to other PKSs (78-85%) among all the constituent domains. Both malonyl-coenzyme A (MCoA) and methylmalonyl-coenzyme A (mMCoA) are substrates for chain elongation steps carried out by the Rf PKS. Since acyltransferase (AT) domains of modular PKSs can distinguish between these two substrates, comparison of the sequence of all ten AT domains of the Rf PKS with those found in the erythromycin (Er) (Donadio, S. and Katz, L., 1992. Gene 111, 51-60) and rapamycin (Rp) (Haydock, S. et al., 1995. FEBS Lett. 374, 246-248) PKSs revealed that the AT domains in module 2 of RifA and module 9 of RifE are specific for MCoA, whereas the other eight modules specify mMCoA. Dehydration of the beta-hydroxyacylthioester intermediates should occur during the reactions catalysed by module 4 of RifB and modules 9 and 10 of RifE, yet only the active site region of module 4 conforms closely to the dehydratase (DH) motifs in the Er and Rp PKSs. The DH domains of modules 9 and 10 diverge significantly from the consensus sequence defined by the Er and Rp PKSs, except for the active site His residues. Deletions in the DH active sites of module 1 in RifA and module 5 in RifB and in the N- and C-terminal regions of module 8 of RifD should inactivate these domains, and module 2 of RifA lacks a DH domain, all of which are consistent with the proposed biosynthesis of Rf. In contrast, module 6 of RifB and module 7 of RifC appear to contain intact DH domains even though DH activity is not apparently required in these modules. Module 2 of RifA lacks a beta-ketoacyl:acyl carrier protein reductase (KR) domain and the one in module 3 has an apparently inactive NADPH binding motif, similar to one found in the Er PKS, while the other eight KR domains of the Rf PKS should be functional. These observations are consistent with biosynthetic predictions. All the acyl carrier protein (ACP) domains, while clearly functional, nevertheless have active site signature sequences distinctive from those of the Er and Rp PKSs. Module 2 of RifA has only the core domains (KS, AT and ACP). The starter unit ligase (SUL) and ACP domains present in the N-terminus of RifA direct the selection and loading of the starter unit, 3-amino-5-hydroxybenzoic acid (AHBA), onto the PKS. AHBA is made by the products of several other genes in the Rf cluster through a variant of the shikimate pathway (August, P.R. et al., inter alia). RifF, produced by the gene immediately downstream of rifE, is thought to catalyse the intramolecular cyclization of the PKS product, thereby forming the ansamacrolide precursor of Rf B. 1998 Elsevier Science B.V.

Interactions between the middle ear and the inner ear: bacterial products.

The round-window membrane (RWM) is extremely thin and is the only soft-tissue barrier between the middle ear and the inner ear. Under inflammatory conditions of the middle ear the various layers of the triple-layered RWM undergo characteristic changes parallel to the changes of the middle-ear mucosa. Several studies report that bacterial products, exo- and endotoxins, from bacteria invading the middle ear may result in profound inflammatory changes in the inner ear, followed by severe damage to the inner-ear function. The present review, in which we summarized experimental and clinical observations, on bacterial products in interactions between the middle and inner ear, focused on: 1. Bacteria and bacterial products in an inflamed middle ear that may influence inner-ear function. 2. RWM structure and RWM permeability under the influence of bacteria and bacterial products. 3. Morphological and functional inner-ear effects of bacterial infection of the middle ear, and the possible mechanisms involved. 4. Future studies to be directed in this field.

Pharmacological characterization of effects of verapamil and GS 283 on isolated guinea pig and rat trachealis.

The pharmacological effects of verapamil and GS 283, 1-(4'-methoxybenzyl)-6,7-dihydroxy-3,4-dihydroxyisoquinoline, were investigated using isolated rat and guinea pig trachealis. Both verapamil and GS 283 inhibited carbachol-, histamine (only guinea pig)-, and high-K(+)-induced contraction in a dose-dependent manner. GS 283 acted as a weak histamine H1 and muscarinic receptor antagonist in guinea pig and rat trachealis with respective pKB values in the range of 5.60 approximately 6.12 and 5.17 approximately 5.83. On the other hand, pyrilamine and atropine showed a typical competitive antagonism on histamine (guinea pig) and on muscarinic receptors (rat trachea) with pKB values of 9.25 +/- 0.21 and 9.37 +/- 0.32, respectively. GS 283 inhibited Ca(2+)-induced contraction on guinea pig trachealis in Ca(2+)-free media. Furthermore, very high concentrations of GS 283 and verapamil completely abolished a phasic contraction induced by carbachol in Ca(2+)-free media, suggesting that verapamil and GS 283 can enter into the cytoplasm, where they may exert secondary actions on internal sites of the muscle, such as the sarcoplasmic reticulum. It is concluded that GS 283 has a Ca2+ antagonistic action along with weak histamine and muscarinic receptor blocking activity in isolated rat and guinea pig tracheal smooth muscle and its mode of action is likely inhibition of Ca2+ influx from plasma membrane and also release from the sarcoplasmic reticulum.

Thioesterases and the premature termination of polyketide chain elongation in rifamycin B biosynthesis by Amycolatopsis mediterranei S699.

The role of two thioesterase genes in the premature release of polyketide synthase intermediates during rifamycin biosynthesis in the Amycolatopsis mediterranei S699 strain was investigated. Creation of an in-frame deletion in the rifR gene led to a 30 approximately 60% decrease in the production of both rifamycin B by the S699 strain or a series of tetra- to decaketide shunt products of polyketide chain assembly by the rifF strain. Since a similar percentage decrease was seen in both genetic backgrounds, we conclude that the RifR thioesterase 2 is not involved in premature release of the carbon chain assembly intermediates. Similarly, fusion of the Saccharopolyspora erythraea DEBS3 thioesterase I domain to the C-terminus of the RifE PKS subunit did not result in a noticeable increase in the amount of the undecaketide intermediate formed nor in the amounts of the tetra- to decaketide shunt products. Hence, premature release of the carbon chain assembly intermediates is an unusual property of the Rif PKS itself.

Estimation of the effective transversely isotropic elastic constants of a material from known values of the material's orthotropic elastic constants.

A method is illustrated for determining the effective transversely isotropic (or isotropic) elastic constants from measured orthotropic elastic constants. This method consists of constructing upper and lower bounds on the effective transversely isotropic (or isotropic) elastic constants using the known orthotropic values. This method is illustrated using three sets of elastic constants for bone. Fortunately, the upper and lower bounds are very close. Thus very good approximations for the effective transversely isotropic (or isotropic) elastic constants for cortical and cancellous bone are obtained from previously published data on the orthotropic elastic constants for those tissue types. This work is undertaken to build a greater database for the transversely isotropic elastic constants of bone with the intention of employing them in a transversely isotropic model of bone poroelasticity. An interesting aspect of the present result is that the Voigt and Reuss bounds are very tight for these anisotropic materials. This is not always the case for these bounds.

Immunohistochemical localization of alpha-atrial natriuretic polypeptide in the rat cochlea.

The localization of alpha-atrial natriuretic polypeptide (alpha-ANP) in the rat cochlea was studied by immunohistochemical technique, using a polyclonal antibody against synthetic rat alpha-atrial natriuretic polypeptide (alpha-rANP). The spiral ligament of the lateral cochlear wall exhibited pronounced immunoreactivity to atrial natriuretic polypeptide (ANP), whereas the stria vascularis displayed almost no immunoreaction. ANP immunoreactivity (IR) was also intense in the spiral limbus region. IR was observed in the fibroblast-like cells and in the extracellular matrix, in particular along its fibrous bundles, of both the spiral ligament and the spiral limbus. These results indicate that ANP may participate in the regulation of the water electrolyte balance at these sites, which may imply a role for ANP in the regulation of cochlear fluids.

Sound field calculations for an ultrasonic linear phased array with a spherical liquid lens.

Lenses are often used to provide focusing in the elevation dimension of ultrasonic linear phased-array transducers. The use of a liquid lens in this application adds a variable geometric focusing capability, determined by the radius of curvature of the lens surface and speed of sound in the liquid, to the electronic focusing produced by the linear phased array. An efficient method to calculate the sound field radiated from the linear phased-array transducer through the liquid lens is presented. It treats the lens surface as a secondary source distribution according to Huygens's principle, and employs a modified form of the rectangular radiator method to calculate the field. The appropriate phases for the array elements to focus and steer the beam are calculated by considering the refraction on the lens surface. Comparisons of computer simulations and experimental measurements of the field intensity distribution of a prototype linear array transducer with a liquid lens demonstrate the accuracy of the proposed method.

Ultrasonic phased arrays with variable geometric focusing for hyperthermia applications.

An ultrasonic applicator, which utilizes both electronic and variable geometric focusing, for deep-localized hyperthermia is investigated. The applicator is based around a linear phased array that furnishes its electronic focusing capability. The output of the array radiates through a spherical liquid-lens that provides the applicator a variable geometric focusing capability as well. A lens of this type adds dynamic focusing to the elevation dimension of the linear phased array. By controlling the volume of liquid in the lens (and thus the radius of curvature of its membrane), dynamic control of the geometrical focus can be achieved. Comparisons of computer simulations and experimental measurements of the field intensity distribution of a small-scale prototype applicator are presented. Important design parameters, such as the choice of the liquid for the lens and the size and number of array elements, are examined.

Ultrasonic phased array controller for hyperthermia applications.

Multiple and mechanically scanned ultrasound transducer systems have demonstrated the efficacy of using ultrasound to produce deep localized hyperthermia. The use of ultrasonic phased arrays has been proposed as an alternative to these systems. A phased array offers a more flexible approach to heating tumours in that the size, shape, and position of its focal region can be altered during the course of treatment in order to achieve the desired temperature distribution. This added flexibility comes at the cost of increased complexity of the hardware necessary to drive the transducer because each element requires its own amplifer with both phase and amplitude control. In order for phased arrays with large numbers of elements to be feasible for hyperthermia applications, the complexity of this circuitry must be minimized. This paper describes a circuit design which simplifies the electronics required to control a phased array transducer system for hyperthermia applications. The design is capable of controlling virtually any type of phased array transducer operating at frequencies less than 2 MHz. The system performance was verified through beam profile measurements using a 48-element tapered phased array transducer.

Effect of head rotation on lateral cephalometric radiographs.

The purpose of this study was to identify the potential projection errors of lateral cephalometric radiographs due to head rotation in the vertical Z-axis. For this investigation, 17 human dry skull samples with permanent dentition were collected from the Department of Anatomy in the College of Medicine, Chosun University. They had no gross asymmetry and were well preserved. Each dry skull was rotated from 0 degrees to +/- 15 degrees at 1 degrees intervals. A vertical axis, the Z-axis, was used as a rotational axis to have 527 lateral cephalometric radiographs exposed. The findings were that: (1) angular measurements have fewer projection errors than linear measurements; (2) the greater the number of landmarks on the midsagittal plane that are included in angular measurements, the fewer the projection errors occurring; (3) horizontal linear measurements decrease gradually in length as the rotational angle toward the film increases, whereas a small increase and then decrease of the length occurs as the rotational angle toward the focal spot increases; (4) horizontal linear measurements have more projection errors than vertical linear measurements according to head rotation; and (5) projection errors of vertical linear measurements increase as the distance from the rotational axis increases. In summary, angular measurements of lateral cephalometric radiographs are more useful than linear measurements in minimizing the projection errors associated with head rotation on a vertical axis.