Evaluating a combination treatment of NK cells and reovirus against bladder cancer cells using an in vitro assay to simulate intravesical therapy.

Intravesical treatment using either reovirus or natural killer (NK) cells serves as an efficient strategy for the treatment of bladder cancer cells (BCCs); however, corresponding monotherapies have often shown modest cytotoxicity. The potential of a locoregional combination using high-dose reovirus and NK cell therapy in an intravesical approach has not yet been studied. In this study, we evaluated the effectiveness of reoviruses and expanded NK cells (eNK) as potential strategies for the treatment of bladder cancer. The anti-tumor effects of mono-treatment with reovirus type 3 Dearing strain (RC402 and RP116) and in combination with interleukin (IL)-18/-21-pretreated eNK cells were investigated on BCC lines (5637, HT-1376, and 253J-BV) using intravesical therapy to simulate in vitro model. RP116 and IL-18/-21-pretreated eNK cells exhibited effective cytotoxicity against grade 1 carcinoma (5637 cells) when used alone, but not against HT-1376 (grade 2 carcinoma) and 253J-BV cells (derived from a metastatic site). Notably, combining RP116 with IL-18/-21-pretreated eNK cells displayed effective cytotoxicity against both HT-1376 and 253J-BV cells. Our findings underscore the potential of a combination therapy using reoviruses and NK cells as a promising strategy for treating bladder cancer.

The pathophysiology of visceral adipose tissues in cardiometabolic diseases.

Central pattern of fat distribution, especially fat accumulation within the intraabdominal cavity increases risks for cardiometabolic diseases. Portal hypothesis combined with a pathological remodeling in visceral fat is considered the major etiological factor explaining the independent contribution of visceral obesity to cardiometabolic diseases. Excessive remodeling in visceral fat during development of obesity leads to dysfunctions in the depot, characterized by hypertrophy and death of adipocytes, hypoxia, inflammation, and fibrosis. Dysfunctional visceral fat secretes elevated levels of fatty acids, glycerol, and proinflammatory and profibrotic cytokines into the portal vein directly impacting the liver, the central regulator of systemic metabolism. These metabolic and endocrine products induce ectopic fat accumulation, insulin resistance, inflammation, and fibrosis in the liver, which in turn causes or exacerbates systemic metabolic derangements. Elucidation of underlying mechanisms that lead to the pathological remodeling and higher degree of dysfunctions in visceral adipose tissue is therefore, critical for the development of therapeutics to prevent deleterious sequelae in obesity. We review depot differences in metabolic and endocrine properties and expendabilities as well as underlying mechanisms that contribute to the pathophysiological aspects of visceral adiposity in cardiometabolic diseases. We also discuss impacts of different weight loss interventions on visceral adiposity and cardiometabolic diseases.

Metabolic and structural remodeling during browning of primary human adipocytes derived from omental and subcutaneous depots.

OBJECTIVE: This study investigated remodeling of cellular metabolism and structures during browning of primary human adipocytes derived from both visceral and subcutaneous adipose tissues. Effects of glucocorticoids on the browning were also assessed. METHODS: Differentiated omental and subcutaneous human adipocytes were treated with rosiglitazone, with or without dexamethasone, and expression levels of brite adipocyte markers, lipolysis, and lipid droplet and mitochondrial structures were examined. RESULTS: Both omental and subcutaneous adipocytes acquired brite phenotypes upon peroxisome proliferator-activated receptor-γ agonist treatment, and dexamethasone tended to enhance the remodeling. Although rosiglitazone increased lipolysis during treatment, brite adipocytes exhibited lower basal lipolytic rates and enhanced responses to ß-adrenergic agonists or atrial natriuretic peptide. Transcriptome analysis identified induction of both breakdown and biosynthesis of lipids in brite adipocytes. After 60+ days in culture, lipid droplet size increased to ~50 microns, becoming almost unilocular in control adipocytes, and after browning, they acquired paucilocular morphology, clusters of small lipid droplets (1-2 micron) surrounded by mitochondria appearing on the periphery of the central large one. CONCLUSIONS: Metabolic and structural remodeling during browning of primary human adipocytes is similar to previous findings in human adipocytes in vivo, supporting their uses for mechanical studies investigating browning with translational relevance.

Evaluating the Suitability of the Plantaris Tendon for Sports Trauma Reconstruction and a Predictive Model of Tendon Length Based on Height and Leg Length.

This study evaluates the suitability of the plantaris tendon (PT) as a tendon graft donor for sports trauma reconstruction and proposes a predictive model for estimating PT length by using an individual's height and leg length. Anatomical dissection of 50 cadavers (32 males and 18 females) yielded precise measurements of PT length and width while also recording height and leg length. Among the lower limbs, 89% were suitable for at least one recommended graft suitability criterion. In addition, PT length exhibited robust positive correlations with height and leg length. Predictive equations were established for estimating the PT length based on leg length and height with consistency across sexes and sides: PT length = 0.605 + 0.396 × leg length (r = 0.721) and PT length = 1.480 + 0.193 × height (r = 0.626). This study underscores the grafting potential of the PT, providing a predictive tool that can aid surgeons in addressing tendon graft challenges within sports trauma scenarios.

Fenofibrate alleviates insulin resistance by reducing tissue inflammation in obese ovariectomized mice.

BACKGROUND: Fenofibrate is a hypolipidemic peroxisome proliferator-activated receptor α (PPARα) agonist used clinically to reduce hypercholesterolemia and hypertriglyceridemia. OBJECTIVE: We investigated the effects of fenofibrate on insulin resistance and tissue inflammation in a high-fat diet (HFD)-fed ovariectomized (OVX) C57BL/6J mice, a mouse model of obese postmenopausal women. METHODS: Female OVX mice were randomly divided into 3 groups and received a low-fat diet, an HFD, or an HFD supplemented with 0.05% (w/w) fenofibrate for 9 weeks. Parameters of insulin resistance and tissue inflammation were measured using blood analysis, histological analysis, immunohistochemistry, and quantitative real-time polymerase chain reaction. RESULTS: When fenofibrate was administered to HFD-fed OVX mice for 9 weeks, we observed reductions in body weight gain, adipose tissue mass, and the size of visceral adipocytes without the change of food intake. Fenofibrate improved mild hyperglycemia, severe hyperinsulinemia, and glucose tolerance in these mice. It also reduced pancreatic islet size and insulin-positive ß-cell area to levels similar to those in OVX mice fed a low-fat diet. Concomitantly, administration of fenofibrate not only suppressed pancreatic lipid accumulation but also decreased CD68-positive macrophages in both the pancreas and visceral adipose tissue. Treatment with fenofibrate reduced tumor necrosis factor α (TNFα) mRNA levels in adipose tissue and lowered serum TNFα levels. CONCLUSION: These results suggest that fenofibrate treatment attenuates insulin resistance in part by reducing tissue inflammation and TNFα expression in HFD-fed OVX mice.

l-Arginine supplementation regulates energy-substrate metabolism in skeletal muscle and adipose tissue of diet-induced obese rats.

Dietary supplementation with l-arginine has been reported to reduce white fat mass in diet-induced obese rats and in obese humans. This study was conducted to test the hypothesis that the arginine treatment regulates glucose and fatty acid metabolism in insulin-sensitive tissues. Male Sprague-Dawley rats (4-week-old) were fed either low- or high-fat diets for 15 weeks (n = 16/diet). Thereafter, lean or obese rats were fed their respective diets and received drinking water containing either 1.51% l-arginine-HCl or 2.55% alanine (isonitrogenous control) (n = 8/treatment group). After 12 weeks of treatment, rats were euthanized and tissue samples were collected for biochemical assays. High-fat feeding increased the size of adipocytes isolated from retroperitoneal (RP) adipose tissue, while arginine treatment reduced their size. The total number of adipocytes in the adipose tissue did not differ among the four groups of rats. Glucose oxidation in extensor digitorum longus (EDL) muscle, soleus muscle, and RP adipose tissue were reduced in response to high-fat feeding. On the contrary, oleic acid oxidation in RP adipose tissue was enhanced in rats fed the high-fat diet. Arginine treatment stimulated both glucose and oleic acid oxidation in EDL and soleus muscles, while having no effect on glucose oxidation, oleic acid oxidation, or basal lipolysis per 106 adipocytes in RP adipose tissue. Collectively, these results indicate that oral supplementation with arginine to diet-induced obese rats promoted the oxidation of energy substrates in skeletal muscle, thereby reducing white fat in the body.

Remodeling of Adipose Tissues by Fatty Acids: Mechanistic Update on Browning and Thermogenesis by n-3 Polyunsaturated Fatty Acids.

Enhancing thermogenesis by increasing the amount and activity of brown and brite adipocytes is a potential therapeutic target for obesity and its associated diseases. Diet plays important roles in energy metabolism and a myriad of dietary components including lipids are known to regulate thermogenesis through recruitment and activation of brown and brite adipocytes. Depending on types of fatty acids (FAs), the major constituent in lipids, their health benefits differ. Long-chain polyunsaturated FAs (PUFAs), especially n-3 PUFAs remodel adipose tissues in a healthier manner with reduced inflammation and enhanced thermogenesis, while saturated FAs exhibit contrasting effects. Lipid mediators derived from FAs act as autocrine/paracrine as well as endocrine factors to regulate thermogenesis. We discuss lipid mediators that may contribute to the differential effects of FAs on adipose tissue remodeling and hence, cardiometabolic diseases. We also discuss current understanding of molecular and cellular mechanisms through which n-3 PUFAs enhance thermogenesis. Elucidating molecular details of beneficial effects of n-3 PUFAs on thermogenesis is expected to provide information that can be used for development of novel therapeutics for obesity and its associated diseases.

Normal Distal Tibiofibular Syndesmosis Assessment Using Postmortem Computed Tomography (PMCT).

BACKGROUND: Distal tibiofibular syndesmotic injuries, often misdiagnosed, can lead to substantial morbidity. This study utilized postmortem computed tomography (PMCT) to define normal syndesmotic relationships in 131 subjects. METHODS: Three parameters were measured: fibular rotation (FR), sagittal translation (ST), and incisura depth (ID). RESULTS: Interobserver reliability was excellent for FR and ID but moderate for ST. Anatomical variability was wide, with FR ranging from -0.4° to 16.6°, ST from 0.33 mm to 3.49 mm, and ID from 1.89 mm to 6.05 mm. Side-to-side variability within subjects was minimal. Gender-specific differences were observed in ST, possibly due to size variations, highlighting the need for gender-specific diagnostic criteria. CONCLUSIONS: Although establishing universal reference values is challenging, using contralateral ankles for comparison can enhance diagnostic accuracy in syndesmotic injuries. This study, the first of its kind, offers valuable insights into normal distal tibiofibular syndesmotic relationships based on PMCT data. Future validation studies in patients with syndesmotic injuries can further improve diagnostic accuracy.

Evaluation of Barley and Wheat Germplasm for Resistance to Head Blight and Mycotoxin Production by Fusarium asiaticum and F. graminearum.

Fusarium head blight (FHB) is one of the most serious diseases in barley and wheat, as it is usually accompanied by the production of harmful mycotoxins in the grains. To identify FHB-resistant breeding resources, we evaluated 60 elite germplasm accessions of barley (24) and wheat (36) for FHB and mycotoxin accumulation. Assessments were performed in a greenhouse and five heads per accession were inoculated with both Fusarium asiaticum (Fa73, nivalenol producer) and F. graminearum (Fg39, deoxynivalenol producer) strains. While the accessions varied in disease severity and mycotoxin production, four wheat and one barley showed <20% FHB severity repeatedly by both strains. Mycotoxin levels in these accessions ranged up to 3.9 mg/kg. FHB severity was generally higher in barley than in wheat, and Fa73 was more aggressive in both crops than Fg39. Fg39 itself, however, was more aggressive toward wheat and produced more mycotoxin in wheat than in barley. FHB severity by Fa73 and Fg39 were moderately correlated in both crops (r = 0.57/0.60 in barley and 0.42/0.58 in wheat). FHB severity and toxin production were also correlated in both crops, with a stronger correlation for Fa73 (r = 0.42/0.82 in barley, 0.70 in wheat) than for Fg39.

Estimating Adult Stature Using Metatarsal Length in the Korean Population: A Cadaveric Study.

This study aims to propose a regression equation for estimating stature in the Korean population using metatarsal bones from cadavers and to validate the appropriateness of the Korean-specific equation by comparing it to equations from other populations. A total of 81 adult formalin-fixed cadavers (51 males and 30 females) were evaluated. The first and second metatarsal bones' physiological and maximal lengths were measured, and the cadaveric stature of the subjects was determined as the distance from the vertex to the plantar face of the heel. In all measurements, the correlation coefficient between real stature and metatarsal length was statistically significant (p < 0.001). Additionally, both sexes showed a correlation between stature and metatarsal bone length. For unknown sex, M1 (first metatarsal maximal length) showed the strongest association between stature and metatarsal length. The following is the appropriate regression equation: 1172.4913 + 7.3275M1 (R = 0.703). The current equation demonstrated a statistically significant appropriateness for the Korean population when compared to equations for other populations (p < 0.001). In conclusion, we proposed a Korean-specific regression equation for estimating stature using metatarsal length, and this formula may be more appropriate and useful in forensic science for the Korean population.

Anatomical Characteristics of Extensor Hallucis Longus Tendon Variations and Its Clinical Implications: A Korean Cadaveric Study.

The purpose of this study is to ascertain the morphological characteristics of the extensor hallucis longus (EHL) tendon variation using larger-scale dissection of Korean cadavers and to classify the types of variation along with incidence. A total of 158 feet from adult formalin-fixed cadavers (50 males, 29 females) were dissected. The morphological characteristics and measurements of the EHL tendon variants were evaluated. Three types of the EHL tendon variation were classified, wherein the most common type was Type 2 (106 feet, 67.1%), Type 3 (3 feet, 1.9%) was the rarest type, and Type 1 without accessory tendon was found in 49 feet (31.0%). Type 2K (11 feet, 7%) and Type 3K (1 foot, 0.6%) were described as new subtypes. The present study suggests morphological characteristics of the EHL tendon variation in Korean populations and high morphological variability of the EHL tendon along with the possibility of differences according to race or ethnicity and gender. Furthermore, a newly updated classification complemented by new subtypes of variation will help foot and ankle surgeons in diagnosis and surgical planning with hallux problems.

Selective Expansion of NKG2C+ Adaptive NK Cells Using K562 Cells Expressing HLA-E.

Adaptive natural killer (NK) cells expressing self-specific inhibitory killer-cell immunoglobulin-like receptors (KIRs) can be expanded in vivo in response to human cytomegalovirus (HCMV) infection. Developing a method to preferentially expand this subset is essential for effective targeting of allogeneic cancer cells. A previous study developed an in vitro method to generate single KIR+ NK cells for enhanced targeting of the primary acute lymphoblastic leukemia cells; however, the expansion rate was quite low. Here, we present an effective expansion method using genetically modified K562-HLA-E feeder cells for long-term proliferation of adaptive NK cells displaying highly differentiated phenotype and comparable cytotoxicity, CD107a, and interferon-γ (IFN-γ) production. More importantly, our expansion method achieved more than a 10,000-fold expansion of adaptive NK cells after 6 weeks of culture, providing a high yield of alloreactive NK cells for cell therapy against cancer.

Anatomical Classification for Plantaris Tendon Insertion and Its Clinical Implications: A Cadaveric Study.

The purposes of this study were to ascertain the morphological characteristics of a plantaris tendon (PT) insertion using a larger-scale dissection of Korean cadavers and to classify the types of PT insertion related to the calcaneal tendon (CT). A total of 108 feet from adult formalin-fixed cadavers (34 males, 20 females) were dissected. The morphological characteristics and measurements of the PT insertion were evaluated. Five types of PT insertion were classified, wherein the most common type was Type 1 (39 feet, 63.1%). Type 2 and Type 3 were similar, with 16 feet (14.8%) and 15 feet (13.9%), respectively. Type 4 (6 feet, 5.6%) was the rarest type, and Type 5 had 25 feet (23.1%). The case of an absent PT was noted in 7 feet (6.5%). In the proximal portion, the tendon had a thick and narrow shape, became thin and wide in the middle portion, and then changed to thick and narrow again just before the insertion into the calcaneal tuberosity. This study confirmed the five types according to the location of the PT and the area of its insertion-related CT. The morphology of the PT insertion may be anatomically likely to influence the occurrence of tendinopathy in the CT.

Functional roles of all five putative hydrophobin genes in growth, development, and secondary metabolism in Fusarium graminearum.

Fusarium graminearum is the causal agent of Fusarium head blight in cereal crops. As in other filamentous ascomycetes, F. graminearum contains genes encoding putative hydrophobins, which are small secreted amphiphilic proteins with eight conserved cysteine residues. Here, we investigated the roles of all five hydrophobin genes (designated FgHyd1, FgHyd2, FgHyd3, FgHyd4, and FgHyd5) in various mycological traits of F. graminearum. Gene expression analyses revealed that the five FgHyd genes, all of which were under the control of G protein signaling or velvet complex proteins, were differentially expressed under various developmental conditions. Three genes (FgHyd1, FgHyd2, and FgHyd3) were constitutively expressed in all aerial structures examined (hyphae, conidia, and perithecia), and two genes (FgHyd1 and FgHyd2) were also expressed in submerged hyphae. FgHyd3 was exclusively expressed in aerial hyphae on solid surfaces, including rice grains. These genes showed markedly reduced expression in F. asiaticum, which was a closely related to F. graminearum but exhibited different mycological traits from F. graminearum. Phenotypic analyses of various gene deletion strains, including the quintuple deletion (ΔFgHyd12345) strain, confirmed that in addition to their typical functions, all five FgHyd genes were involved in other traits, such as conidiation, pathogenicity, and secondary metabolism in F. graminearum. Both RNA-seq and chemical analyses confirmed that ΔFgHyd led to overproduction of specific terpenoid compounds (e.g., trichothecenes), which has not been reported previously. Nevertheless, the lack of complete phenotypic loss of any of the traits examined, even in the ΔFgHyd12345 strain, and little cumulative action of all five FgHyd genes strongly suggest that all five hydrophobins are redundant in function and are not absolutely essential for these fungal traits in F. graminearum.

Effect of Steam and Smoke Cooking Processes on Web-Foot Octopus (Amphioctopus sp.) Home Meal Replacement Product.

In Korea, the web-foot octopus (Amphioctopus sp.) is commonly consumed as jjukkumi bokkeum, a spicy stir-fried octopus dish. Using steaming and smoking methods, we made jjukkumi bokkeum home meal replacement (HMR) products. The response surface methodology (RSM) was employed to optimize the steam and smoke processes. Quick freezing was applied to freeze the test product at -35 °C. Then, the physicochemical, biological, nutritional characteristics, and shelf-life of the test HMR products were evaluated. The optimal conditions for steaming and smoking were 95 °C for 2 min and 70 °C for 11 min, respectively. The pH, volatile basic nitrogen content, and thiobarbituric acid-reactive substances content decreased after steaming and smoking, indicating that these processes maintained these parameters well. Sensory evaluation revealed that there were no changes in these characteristics after freezing and reheating. Further, the test HMR products contained the daily nutritional requirements of macro and micronutrients, as well as amino acids and fatty acids. The shelf-life of the HMR products was estimated to be 15 months. The findings of this study indicate that the application of steam and smoke processes to produce a jjukkumi bokkeum HMR product results in a high-quality product with a long shelf-life.

Adult Stature Estimation from Radiographic Metatarsal Length in a Contemporary Korean Population.

The ability to estimate stature can be important in the identification of skeletal remains. This study aims to develop a Korean-specific equation predicting stature using radiographic measurements in the contemporary Korean population. 200 healthy Korean adults, including 102 males and 98 females, were randomly selected (age, range 20-86 years). The first and second metatarsals of the foot were measured by a standing X-ray using a digital medical image viewer. The result showed a statistically significant correlation between metatarsal length and stature in Korean populations (male, R = 0.46, p < 0.001; female, R = 0.454, p < 0.001). Values of correlation coefficients (R) of the equations were 0.431 to 0.477. Compared to equations derived from other races, the Korean-specific equation showed significantly lower error values for estimating the actual height of Koreans through cross-validation. In conclusion, this study is the first to propose a Korean-specific regression formula for estimating stature using metatarsal length and a verified formula for precise application to the Korean population. However, given the relatively low correlation coefficient, the stature estimation formula derived from this study can be utilized when other bones that allow more accurate stature estimation are not available.

A New Anatomical Classification for Tibialis Posterior Tendon Insertion and Its Clinical Implications: A Cadaveric Study.

The variations in the tibialis posterior tendon (TPT) could not be defined by previous classification; thus, this study used a larger-scale cadaver with the aim to classify the types of TPT insertion based on the combination of the number and location of TPT insertions. A total of 118 feet from adult formalin-fixed cadavers were dissected (68 males, 50 females). The morphological characteristics and measurements of TPT insertion were evaluated. Four types of TPT insertions were classified, wherein the most common type was type 4 (quadruple insertions, 78 feet, 66.1%), which was divided into four new subtypes that were not defined in the previous classification. The second most common type was type 3 (triple insertions, 25 feet, 21.2%) with three subtypes, including the new subtype. Type 2 was found in 13 feet (11%), and the rarest type was type 1 (2 feet, 1.7%), wherein the main tendon was only attached to the navicular bone and the medial cuneiform bone. We suggest high morphological variability of the TPT in relation to the insertion location, along with the possibility of significant differences according to race and gender. Moreover, this classification will help clinicians understand adult flatfoot deformity-related posterior tibial tendon dysfunction (PTTD).

Acetylated Resveratrol and Oxyresveratrol Suppress UVB-Induced MMP-1 Expression in Human Dermal Fibroblasts.

Resveratrol (RES) and oxyresveratrol (OXYRES) are considered and utilized as active ingredients of anti-aging skin cosmetics. However, these compounds are susceptible to oxidative discoloration and unpleasant odor in solutions, limiting their use in cosmetics. Accordingly, RES and OXYRES were chemically modified to acetylated derivatives with enhanced stability, and their anti-aging effect on the skin and detailed molecular mechanism of their acetylated derivatives were investigated. Acetylated RES and OXYRES lost their acetyl group and exerted an inhibitory effect on H2O2-induced ROS levels in human dermal fibroblast (HDF) cells. In addition, RES, OXYRES, and their acetylated derivatives suppressed UVB-induced matrix metalloproteinase (MMP)-1 expression via inhibition of mitogen-activated protein kinases (MAPKs) and Akt/mTOR signaling pathways. Furthermore, RES, OXYRES, and their acetylated derivatives suppressed type I collagen in TPA-treated HDF cells. Collectively, these results suggest the beneficial effects and underlying molecular mechanisms of RES, OXYRES, and their acetylated derivatives for anti- skin aging applications.

Effects of Vacuum Frying on the Preparation of Ready-to-Heat Batter-Fried and Sauced Chub Mackerel (Scomber japonicus).

Chub mackerel (CM) is a commercial fish in Korea, owing to its availability and nutritional values. This study aimed to develop a ready-to-heat (RTH) Korean preparation of CM, known as Godeungo gangjeong. We utilized vacuum frying technology to fry the CM and evaluated its quality. Conventional frying with a deep fryer was performed in parallel to assess the superiority of the vacuum fryer. We optimized the frying conditions of vacuum frying (VBF) and deep frying (DBF) using response surface methodology. At optimum conditions of 95 °C for 7 min 42 s, VBF produced better sensory, chemical, and microbial properties than DBF at 190 °C for 5 min 30 s. The nutritional values, including amino acid and fatty acid contents, were investigated and found to be higher in VBF than in DBF. Sensory properties also showed better scores on VBF than DBF, especially in appearance, aroma, taste, and overall acceptability. The VBF produced lower volatile basic nitrogen (VBN), thiobarbituric acid reactive substances (TBARS), and total bacterial count (TBC) than DBF. The findings confirmed that vacuum frying is a better option to produce RTH Godeungo gangjeong, since it provides less oxidation and maintains the product quality. Using the Arrhenius approach, the product was concluded to preserve both quality and safety for 9 months of storage at -18 °C.

The Location of the Fibular Tunnel for Anatomically Accurate Reconstruction of the Lateral Ankle Ligament: A Cadaveric Study.

We aimed to describe the location of fibular footprint of each anterior talofibular ligament (ATFL) and calcaneofibular ligament (CFL), as well as their common origin in relation to bony landmarks of the fibula in order to determine the location of the fibular tunnel. In 105 ankle specimens, the center of the footprints of the ATFL and CFL (cATFL and cCFL, respectively) and the intersection point of their origin (intATFL-CFL) were investigated, and the distances from selected bony landmarks (the articular tip (AT) and the inferior tip (IT) of the fibula) were measured. Forty-two (40%) specimens had single-bundle ATFL, and 63 (60%) had double-bundle patterns. The distance between intATFL-CFL and IT was 12.0 ± 2.5 mm, and a significant difference was observed between the two groups (p = 0.001). Moreover, the ratio of the intATFL-CFL location based on the anterior fibular border for all cadavers was 0.386. The present study suggests a reference ratio that can help surgeons locate the fibular tunnel for a more anatomically accurate reconstruction of the lateral ankle ligament. Also, it may be necessary to make a difference in the location of the fibular tunnel according to the number of ATFL bundles during surgery.