OBJECTIVE: Traditional lubricants can interfere with sperm function. Fertility lubricants have been developed to not harm sperm. This study compared sperm function, including the ability to support embryo development, following contact with glycerol-free (GF) versus glycerol-containing (GC) fertility lubricants.DESIGN: Prospective, controlled.MATERIALS AND METHODS: Two experiments were done to evaluate sperm function after 30 min incubations in control media without lubricant or in treatments of media and 10% v/v lubricant including: (1) GF Pre∼Seed® (INGfertility, Spokane, WA); (2) GC ConceiveEase® (Sepal, Boston, MA); (3) GC Conceive Plus® Lot 19050; and (4) GC Conceive Plus® Lot 199X (different formulations by Sasmar, Brussels, BE). In Experiment 1, human donor sperm (n=10) were cultured in human tubal fluid medium or treatments and subjective motility evaluated. In Experiment 2, cryopreserved bull sperm were cultured in Tyrode's medium or treatments and placed with bovine oocytes (n= 40+) for IVF. Fertilization at 32 hrs and development to morulae on Day 7 were compared using Friedman's.Tabled 1Outcomes Following Sperm Exposure to LubricantsTreatment% Fertilization% Normal Embryos1102 (8)104 (18)292 (21)*59 (22)**378 (5)**49 (15)**473 (4)**67 (3)**Means (SE) as percent of control, differ at *p=0.015, **p<0.001. Open table in a new tab CONCLUSION: Decreased ability of sperm to fertilize and support embryo development after contact with GC lubricants suggested sublethal damage had occured. Bovine and human embryos share paternal regulation patterns, making the model valuable for detection of sublethal sperm damage impacting embryo outcomes. Maintenance of sperm function after contact with the GF lubricant supports its potential use in fertility medicine. OBJECTIVE: Traditional lubricants can interfere with sperm function. Fertility lubricants have been developed to not harm sperm. This study compared sperm function, including the ability to support embryo development, following contact with glycerol-free (GF) versus glycerol-containing (GC) fertility lubricants. DESIGN: Prospective, controlled. MATERIALS AND METHODS: Two experiments were done to evaluate sperm function after 30 min incubations in control media without lubricant or in treatments of media and 10% v/v lubricant including: (1) GF Pre∼Seed® (INGfertility, Spokane, WA); (2) GC ConceiveEase® (Sepal, Boston, MA); (3) GC Conceive Plus® Lot 19050; and (4) GC Conceive Plus® Lot 199X (different formulations by Sasmar, Brussels, BE). In Experiment 1, human donor sperm (n=10) were cultured in human tubal fluid medium or treatments and subjective motility evaluated. In Experiment 2, cryopreserved bull sperm were cultured in Tyrode's medium or treatments and placed with bovine oocytes (n= 40+) for IVF. Fertilization at 32 hrs and development to morulae on Day 7 were compared using Friedman's. Means (SE) as percent of control, differ at *p=0.015, **p<0.001. CONCLUSION: Decreased ability of sperm to fertilize and support embryo development after contact with GC lubricants suggested sublethal damage had occured. Bovine and human embryos share paternal regulation patterns, making the model valuable for detection of sublethal sperm damage impacting embryo outcomes. Maintenance of sperm function after contact with the GF lubricant supports its potential use in fertility medicine.

In all mammalian species studied to date, sperm cells are arrested in the caudal isthmus before ovulation to form an oviductal reservoir. When ovulation occurs, discrete numbers of sperm are released from this region and ascend to the ampulla where fertilization normally occurs. Experimental evidence indicates that the sperm reservoir is instrumental in delivering the appropriate number of sperm in the correct physiological state for normal fertilization. In this paper, we present a comprehensive review of the sperm reservoir formation and functions in mammals. Sperm reservoir manipulation as a potential tool to improve reproductive performance is also discussed

A gene-based approach was used to annotate 322 168 cattle expressed sequence tags (ESTs) based on human genes in order to census the transcriptomes, analyze their expression similarities, and identify genes preferentially expressed in different bovine tissues and (or) organs. Of the 34 157 human coding genes used in a standalone BLAST search, 14 928 could be matched with provisional orthologous sequences in a total of 230 135 bovine ESTs. The remaining 92 033 bovine ESTs were estimated to represent an additional 5970 genes in cattle. On average, ~8600 genes were estimated to be expressed in a single tissue and (or) organ and 13 000 in a pooled tissue library. On the basis of the estimated numbers of genes, no more than 3% of genes would be missed when ~34 000 ESTs were sequenced from a single tissue and (or) organ library and ~40 000 ESTs from a pooled source, respectively. Cluster analyses of the gene expression patterns among 12 single tissues and (or) organs in cattle revealed that their expression similarities would depend on physiological functions. In addition, a total of 1502 genes were identified as preferentially expressed genes in these 12 single tissues and (or) organs with LOD (logarithm of the odds, base 10) ≥ 3.0. Therefore, our study provides some insights for further investigating the developmental and functional relations of various tissues and organs in mammals.Key words: cattle, expressed sequence tags (ESTs), orthologous genes, comparative gene-based approach, in silico census, tissue/organs, transcriptomes.

A comparative gene-based approach was developed to annotate the public porcine ESTs and identify gene expression patterns in embryos and reproductive tissues. This gene-based strategy is very straightforward: the human gene sequences were used as references to accurately catalogue and characterize the gene encoded in livestock species by assembling all ESTs that could be orthologous. Of 33,308 human genes studied, 14,161 were shown to have orthologous EST sequences in pigs. Embryos and reproductive tissues preferentially expressed 724 and 312 genes, respectively. Of these, 22 and 20 were abundantly expressed as basic active genes in embryos and reproductive tissues based on the cut-off criteria. In silico analysis is able to reveal not only previously well-defined genes, but also previously undefined genes preferentially expressed in different tissues. The study provides a panel of transcripts in a broad sense that can be used as targets to study the mechanisms involved in embryo development and reproduction in pigs as well as in other mammals, including humans.

The success of assisted reproductive techniques, such as IVF, could be enhanced by being able to select the most competent spermatozoa in a sample. Attachment and subsequent release of spermatozoa from oviductal epithelial cells (OEC) could provide populations of functionally superior spermatozoa for use in these protocols. The objective of the present study was to investigate the ability of heparin and Ca2+-free medium to induce spermatozoa release from bovine OEC. Epithelial cells were grown to confluence in 24-well plates and pooled frozen bull semen was added to a final concentration of 1 × 106 spermatozoa/well. Spermatozoa were allowed to bind to OEC for 2 h. Medium with unbound spermatozoa was removed and replaced by Sperm-TALP, only (control), with heparin (5, 10, or 15 IU/mL), or Ca2+-free with 2 mM EGTA. Treatments were left on sperm-OEC co-cultures for 0.5, 1, 2, 3, or 5 h. At each time, the media were recovered and spermatozoa from each treatment were counted and evaluated for acrosome integrity and motility. The total number of spermatozoa attached to OEC after 2 h of co-culture was considered 100%. Spermatozoa release is expressed as percentage of the total number of sperm cells bound to OEC after 2 h of co-culture. Data were analyzed by ANOVA and results are expressed as mean ± SEM from three independent replicates. Beginning at 0.5 h, more sperm cells (P < 0.05) were released from OEC in the heparin groups (10 and 15 IU/mL, 77.3 ± 6.2% and 84.0 ± 6.2%, respectively) as compared to the control (46.4 ± 6.2%). The Ca2+-free medium also induced spermatozoa release when compared with the control, but the effect was not significant until 3 h (38.2 ± 1.9% vs 59.5 ± 6.9%; P < 0.05). The percentage of acrosome reacted spermatozoa was not affected by heparin treatment. Heparin at 10 IU/mL increased (P < 0.05) the percentage of motile spermatozoa, whereas Ca2+-free medium caused the opposite effect at 0.5 h after addition of treatments. We conclude that both heparin and Ca2+-free medium are able to promote spermatozoa displacement from OEC attachment. Based on motility and acrosome status data, we predict that released sperm cells may be used for IVF and other assisted reproductive techniques.

The objective of this experiment was to evaluate the effects of various Wagyu sires on carcass quality traits and palatability attributes of cooked ribeye steaks. Wagyu sires were used and grouped as Old (n = 6) or New (n = 2) sires, based on the chronological order in which they were imported into the United States. One hundred thirteen F1 heifer and steer calves sired by Wagyu bulls out of three different sources of cows were fed a backgrounding diet for 112 d consisting of an 80:20 ratio of roughage:concentrate then grazed on a mixture of orchardgrass and bluegrass pasture for 84 d and finished on a 10:90 ratio of roughage:concentrate diet for 231 d in a feedlot. Progeny from New sires had larger (P < .05) ribeye areas, higher (P < .05) marbling scores, and lower (P < .05) maturity scores than progeny from Old sires. Marbling was positively correlated (P < .05) to brightness (r = .56), texture (r = .60), and fat luster (r = .38). Progeny of New sires had lower shear force values (P < .05) than progeny of Old sires. These results indicate the superiority of New Wagyu sires to produce progeny with more marbling, lower shear force values, and larger ribeye areas than Old Wagyu sires. Furthermore, there are substantial differences between Wagyu sires for carcass quality traits and palatability attributes.

OBJECTIVE:To compare function of cultured cryopreserved stallion spermatozoa in a modified Tyrode's medium (TM), with or without bovine serum albumin (BSA), or in uterine tube (oviduct) epithelial cell (OEC) coculture in TM, with or without BSA.SAMPLE POPULATION:Cryopreserved spermatozoa from 6 proven stallions and OEC from bovine reproductive tracts in follicular phase.PROCEDURE:Thawed spermatozoa were cultured in TM, with or without BSA, or cocultured with OEC monolayers in TM, with or without BSA. Percentages of capacitated and acrosome-reacted spermatozoa were measured at 5 hours for TM cultures. Spermatozoal survival and motility characteristics were observed over time for all culture methods. Number of spermatozoa attaching to OEC were compared for cocultures.RESULTS:Use of TM without BSA altered spermatozoal function in cell-free medium culture and OEC coculture. A higher percentage of spermatozoa were acrosome reacted in TM with BSA, although percentages of capacitated spermatozoa did not differ. Spermatozoa survived longer and maintained superior motion in TM culture without BSA and in OEC cocultures. More spermatozoa were able to attach to OEC in TM without BSA.CONCLUSIONS:Incubation of cryopreserved spermatozoa in media with BSA resulted in rapid decrease in percentage of intact, motile spermatozoa and limited their ability to interact with OEC.CLINICAL RELEVANCE:Current culture media used for assisted reproduction techniques in horses do not provide functionally capacitated spermatozoa. Removal of BSA from such media improves spermatozoal quality and survival.

Experiments were designed to compare rates of embryonic development following oocyte exposure to cyropreserved spermatozoa from bulls of varying proven fertility, utilizing 3 different sperm preparation methods prior to oocyte introduction. These included 1) sperm co-culture with bovine oviductal epithelial cells (BOEC); 2) sperm co-culture with buffalo rat liver cells (BRLC); or 3) control culture in a routine, cell-free culture system. Semen from 9 bulls was classified by lifetime 60- to 90-d nonreturn rates as having either (mean ± SEM) high (n=3) 73.2 ± 3a, medium (n=3) 70.3 ± 2b or low (n=3) 65.8 ± 3c field fertility (acP< 0.01; bcP< 0.05). There was no difference in embryo cleavage rates for spermatozoa from the high (58 ± 18%), medium (57 ± 23%) or low (57 ± 18%) fertility groups. Development to morula or beyond of oocytes fertilized with high (53 ± 30%) or low (58 ± 27%) fertility semen tended (P<0.10) to be higher than of those fertilized with medium fertility (33 ± 28%) semen. This lack of relationship between in vivo fertility and in vitro embryo outcome was consistent across all sperm preparation methods. Therefore, pooled data were used to evaluate the effect of sperm preparation on embryo outcome. There was no difference in embryo cleavage rates between BOEC monolayers (51 ± 22%), BRLC monolayers (60 ± 20%) and the cell-free controls (60 ± 17%). Subsequent embryonic development to compact morula and beyond was higher (P<0.01) with the BRLC monolayer treatment (61 ± 28%) than with the BOEC monolayers (42 ± 33%) or control culture (39 ± 24%). In conclusion, these studies suggest that there is no predictive relationship between bull field fertility (in the ranges evaluated here) and in vitro embryo cleavage or development rates. However, oocytes inseminated with sperm cells co-cultured on BRLC monolayers develop to the morula stage or beyond at a higher rate than oocytes inseminated with spermatozoa from the BOEC or cell-free system.

Formation of a spermatozoa (`sperm') reservoir in the mare is thought to occur through lectin-mediated sperm attachment to the oviductal epithelium. Once attached, prefertilization sperm survival is supported by oviductal factors. Cryopreservation of stallion sperm decreases the number of sperm attaching to oviduct epithelial cells (OEC) and the length of time these sperm survive. Quantification of in vitro interactions between sperm and OEC in a co-culture system may provide an assay for functional integrity of cryopreserved or fresh sperm samples. Additionally, superior additives for in vitro handling of stallion sperm may be isolated from OEC secretory products. Experiment 1 compared first service conception (FSC) rates resulting from the use of cryopreserved sperm of seven stallions, with sperm function in co-culture such as attachment to OEC and subsequent survival time. Stallions were grouped by cumulative FSC rates observed over three seasons as having average (44±3%) or high (65±2%) fertility over a total of 217 first services (31±9 per stallion). Samples from stallions in the high fertility group had more (P=0.04) sperm attached to OEC and longer subsequent sperm survival in co-culture (P=0.05) as compared with those from the average fertility group. FSC rates correlated with numbers of sperm attaching to OEC and their survival time in co-culture (r≥0.71). In Experiment 2, the function of cryopreserved stallion sperm was evaluated in culture with OEC secretory products from three different sources. After 5 h of culture, sperm incubated with medium conditioned by bovine OEC which had been `bioactivated' (e.g. previously exposed to sperm in culture) were found to be more (P≤0.05) motile and capacitated as compared to sperm in basal TALP medium alone. Sperm in this conditioned medium also survived longer (P=0.05; 27±5 h vs. 17±4 h) than did those in control medium.

Objective: To determine if sperm attachment to oviduct epithelial cells (OEC) in vitro is selective for higher quality sperm and if the system requires homologous species OEC.Design: Controlled prospective study with outcomes assayed by a technician blind to sperm treatment groups.Setting: An academic research laboratory.Patient(s): Experiment 1: normospermic donors with children ( 4 donors, 7 ejaculates). Experiment 2: cryopreserved donor samples (4 donors).Intervention(s): Semen collection by masturbation after 48 hours of abstinence.Main Outcome Measure(s): Experiment 1: sperm assays of motility, morphology, membrane integrity, and capacitation status. Experiment 2: sperm chromatin (DNA) integrity and condensation.Result(s): Experiment 1: sperm not attaching to OEC had lower motility, more membrane disruptions, and more acrosome reactions than did control sperm. This selectivity was equivalent for sperm in coculture with all OEC types. Experiment 2: sperm attached to OEC had fewer abnormalities in chromatin structure compared with sperm that were not attached.Conclusion(s): Selective attachment of functionally superior sperm to OEC is likely important during sperm reservoir formation in vivo and may be exploitable in vitro as a method to isolate high-quality sperm for clinical procedures. Such a system does not require human origin OEC. (C) 1999 by American Society for Reproductive Medicine.

Formation of a prefertilization sperm reservoir in mammals is thought to occur via sperm cell attachment to fallopian tube or oviduct epithelial cells (OEC). Recent data suggests that such an interaction also occurs for human sperm in the fallopian tube. We have previously validated an in vitro sperm-OEC coculture system utilizing bovine OEC monolayers to study postejaculatory human sperm physiology. This study was done to evaluate aspects of human sperm attachment to OEC in coculture and to determine if such attachment and subsequent sperm survival differ between fresh and cryopreserved human sperm. In experiment 1, aliquots of fresh (n = 4) or cryopreserved sperm (n = 3) from normospermic donors were placed into coculture with OEC monolayers at dilutions ranging from 2 x 10(5) to 15 x 10(6) sperm per well. Numbers of each type of sperm attaching to OEC at each concentration were determined. In experiment 2, fresh and cryopreserved sperm from the same donors (n = 4) were put into OEC coculture to observe numbers attaching and subsequent survival time for each sperm type. Sperm attachment to OEC occurred in a linear, dose-dependent manner for fresh and cryopreserved sperm in experiment 1, both as a function of total sperm numbers and as a function of numbers of motile sperm applied (R2 > or = 0.79). However, cryopreserved sperm attached to the OEC at a slower rate than fresh (as a function of the average increase in the number of sperm attaching per unit increase in the number of sperm applied; P < 0.05), with an overall lower percentage of the total and motile sperm applied attaching to OEC (P < 0.01) for cryopreserved versus fresh sperm. Fewer cryopreserved sperm also attached to the OEC, as compared with fresh sperm, in experiment 2 (P < 0.05), even after correcting for motility differences between the sperm types. Sperm survival time in coculture was also decreased for cryopreserved sperm as compared with fresh sperm (P = 0.005). Understanding the kinetics of sperm and OEC interactions may be useful for developing improved cryopreservation protocols or bioassays of sperm function.

Human sperm function was compared in co-culture with monolayers of oviduct epithelial cells (OEC) from three species, human, macaque and bovine. For all species, freeze-thawed and passaged OEC from females in the periovulatory phase were used. OEC cultured on an extracellular matrix (Matrigel) formed a monolayer which supported human sperm attachment to OEC from all three species. Spermatozoa in co-culture with OEC from all three species showed prolonged survival and improved motility characteristics over those cultured in medium alone. This paper describes an efficient, repeatable co-culture system for human spermatozoa which supports sperm attachment to OEC and subsequently improves sperm function over that seen in control medium cultures. Because the improved sperm function in co-culture did not differ significantly between human and bovine OEC for those attributes studied, it is proposed that bovine OEC could be used as an alternative to human OEC in certain human sperm coculture studies. Follicular phase bovine OEC from reproductively normal donors are far more accessible than their human counterparts, thus making this co-culture system more widely available for the study of human spermatozoa-female tract interactions.

In a two-year experiment, 54 steers sired by seven Wagyu bulls [American Wagyu Association (AWA) sire numbers 331, 384, 388, 411, 429, 433 and 488] and 15 steers sired by two Angus bulls, all out of Angus-Hereford cows, were used to evaluate the effects of sire and breed on carcass characteristics and fatty acid composition. Steers were given ad-libitum access to a high-concentrate diet (15 % alfalfa cubes and 85 % barley supplement) for at least 170 days. Breed and individual sire effects were analysed. Wagyu-sired steers had higher marbling, maturity and quality scores, more estimated kidney, pelvic and heart fat, larger longissimus dorsi muscle areas, lower fat thicknesses and yield grades than Angus-sired steers (p < 0.05). Steers sired by 388, 411 and 433 had lower fat thicknesses than steers sired by Angus, 429 and 488 (p < 0.05). Steers sired by 384 and 388 had higher marbling scores per cm subcutaneous fat than steers sired by Angus, 429 and 488, and lower fat thickness per 100 kg of carcass weight than Angus-sired steers (p < 0.05). For both subcutaneous fat and longissimus dorsi muscle, Wagyu-sired steers had higher (p < 0.05) percentages of 14:0, 14:1, 16:0, 16:1, and lower percentages of 18:0 than Angus-sired steers. The genetic differences in carcass characteristics among Wagyu sires may enable us to select for improved marbling with less fat in the Wagyu breed. Some statistically significant (p < 0.05) but small differences existed in fatty acid profiles between breeds and among sires.

This study was conducted to assess the ability of the follicular fluid derived from large and small follicles to support the in vitro oocyte maturation and subsequent fertilization and developmental capacity. Oocytes were cultured in bovine follicular fluid aspirated from small (SFF; 2–5 mm in diameter), large (LFF; 10 to 20 mm in diameter) follicles and TCM199 as a control under 5% CO2 in air. All maturation media were supplemented with 1 IU ml−1 pregnant mare serum gonadotropin. After 24 h culture, oocytes were fertilized in vitro with frozen–thawed and heparin-treated (10 μg ml−1, 15 min) bull spermatozoa and cultured in TCM199 with bovine oviductal epithelial cells (BOEC) for 7 days. Maturation of bovine oocytes cultured in LFF was inhibited and the low of male pronucleus formation was observed when compared with that of SFF (maturation rate: 69 vs. 78%; P<0.05; male pronucleus formation rate: 58 vs. 80%; P<0.05). Developmental capacity of bovine oocytes cultured in SFF was significantly (P<0.05) higher than that of LFF (15 vs. 5%), but significantly (P<0.05) lower than that of the control. There were no differences in the number of nuclei per blastocyst obtained after each treatment. These results indicate that the inhibitory action of follicular fluid on in vitro maturation, male pronucleus formation and developmental capacity of bovine oocytes is dependent on the developmental stage of the follicles from which fluid was obtained.

Twenty-seven Wagyu-sired steers were fed for 90 (14 steers) or 170 (13 steers) days to study the effects of time on feed on palatability and fatty acid composition, and the effects of post-mortem aging time (2, 4 or 10 days) on palatability. Hot carcass weight, fat thickness, longissimus dorsi muscle area, yield grade, estimated kidney, pelvic and heart fat and maturity score were increased (p < 0.05) by an additional 80 days on the high concentrate feed, but marbling was not changed (p > 0.05). Feeding the high concentrate diet for 170 days increased Warner-Bratzler shear force values (p < 0.05) and tended to decrease tenderness (p > 0.05), flavor intensity and connective tissue scores. For the 90 day feeding group, 4 days of aging improved connective tissue score (p < 0.05) and tended to increase (p > 0.05) tenderness scores and decrease shear force, compared with 2 days of aging. For the 170 day feeding group, 10 days of aging improved (p < 0.05) shear force and all sensory attributes except flavor intensity, compared to 2 days of aging. An additional 80 days on feed decreased (p < 0.05) stearic acid and total saturated fatty acids (SFA) and generally increased (p < 0.05) monounsaturated fatty acids (MUFA), polyunsaturated fatty acids (PUFA), MUFA:SFA, and PUFA:SFA in subcutaneous fat and longissimus dorsi muscle. The cholesterol content of fat and muscle increased (p < 0.05) as time on feed increased. Ninety days on a high concentrate diet was adequate for yearling crossbred Wagyu steers to produce highly acceptable carcasses. The additional 80 days on feed produced little or no overall benefit and the steers became overfinished and less tender. Ten days post-mortem aging improved (p < 0.05) all palatability attributes except flavor intensity.

The production of recombinant pharmaceutical proteins in plants benefits from the low cost of upstream production and the greater scalability of plants compared to fermenter-based systems. Now that manufacturing processes that comply with current good manufacturing practices have been developed, plants can compete with established platforms on equal terms. However, the costs of downstream processing remain high, in part because of the dedicated process steps required to remove plant-specific process-related impurities. We therefore investigated whether the ideal strategy for the chromatographic removal of tobacco host cell proteins can be predicted by quantitative structure–activity relationship (QSAR) modeling to reduce the process development time and overall costs. We identified more than 100 tobacco proteins by mass spectrometry and their structures were reconstructed from X-ray crystallography, nuclear magnetic resonance spectroscopy and/or homology modeling data. The resulting three-dimensional models were used to calculate protein descriptors, and significant descriptors were selected based on recently-published retention data for model proteins to develop QSAR models for protein retention on anion, cation and mixed-mode resins. The predicted protein retention profiles were compared with experimental results using crude tobacco protein extracts. Because of the generic nature of the method, it can easily be transferred to other expression systems such as mammalian cells. The quality of the models and potential improvements are discussed.

The influence of beef source, cooking method, and refrigerated storage time on the contents of volatile lipid oxidation products of Wagyu and domestic sources of beef was determined. Longissimus dorsi muscle from Japanese Wagyu, American Wagyu, Longhorn, Angus, and U.S. Choice carcasses was boiled or roasted and stored for 0 or 3 days at refrigerator temperatures. With storage time, the contents of lipid oxidation products increased significantly (P < 0.05). Interactions between beef source and storage time were significant (P < 0.05) for hexanal, 2-pentylfuran, octanal, nonanal, and other major lipid oxidation products. Beef source was shown to have a significant effect on the content of these compounds following 3 days of storage but not immediately following cooking (0 day). The contents of lipid oxidation products were higher in the Japanese and American Wagyu breeds than in other domestic beef sources. In general, cooking method had only minor influences on the content of lipid oxidation products.

The influence of beef source and cooking method on neutral (NL) and polar (PL) lipid contents, fatty acid profiles, and cholesterol contents of Wagyu and domestic sources of beef was determined. Longissimus dorsi muscle from Japanese Wagyu, American Wagyu, Longhorn, Angus, and U.S. Choice was boiled or roasted. Beef from Wagyu breeds had a significantly higher (P < 0.05) NL content than that of domestic sources. The NL from Japanese Wagyu samples was lower in saturated fatty acid content and higher in monounsaturated fatty acid content than samples from other beef sources. The PL from the Japanese Wagyu had the lowest content of saturated fatty acids, while the PL from the American Wagyu had the highest content of monounsaturated fatty acids and the lowest content of polyunsaturated fatty acids. Cholesterol content was highest for the Wagyu breeds. The contents of total NL and individual fatty acids were significantly (P < 0.05) higher in the roasted beef than boiled beef. Cooking method did not have a significant effect on the PL content, PL fatty acid profiles, and cholesterol content.