Background: According to the international criteria for hereditary non-polyposis colorectal cancer (HNPCC) diagnostics, cancer patients with a family history or early onset of colorectal tumours showing high microsatellite instability (MSI-H) should receive genetic counselling and be offered testing for germline mutations in DNA repair genes, mainly MLH1 and MSH2. Recently, an oncogenic V600E hotspot mutation within BRAF, a kinase encoding gene from the RAS/RAF/MAPK pathway, has been found to be associated with sporadic MSI-H colon cancer, but its association with HNPCC remains to be further clarified. Methods: BRAF-V600E mutations were analysed by automatic sequencing in colorectal cancers from 206 sporadic cases with MSI-H and 111 HNPCC cases with known germline mutations in MLH1 and MSH2. In addition, 45 HNPCC cases showing abnormal immunostaining for MSH2 were also analysed. Results: The BRAF-V600E hotspot mutation was found in 40% (82/206) of the sporadic MSI-H tumours analysed but in none of the 111 tested HNPCC tumours or in the 45 cases showing abnormal MSH2 immunostaining. Conclusions: Detection of the V600E mutation in a colorectal MSI-H tumour argues against the presence of a germline mutation in either the MLH1 or MSH2 gene. Therefore, screening of these mismatch repair (MMR) genes can be avoided in cases positive for V600E if no other significant evidence, such as fulfilment of the strict Amsterdam criteria, suggests MMR associated HNPCC. In this context, mutation analysis of the BRAF hotspot is a reliable, fast, and low cost strategy which simplifies genetic testing for HNPCC.

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Nuclear transfer stem cells hold considerable promise in the field of regenerative medicine and cell-based drug discovery. In this study, a total of 29 oocytes were obtained from three young (20 -24 years old) reproductive egg donors who had been successful in previous cycles. These oocytes, deemed by intended parents to be in excess of their reproductive needs, were donated for research without financial compensation by both the egg donor and intended parents after receiving informed consent. All intended parents successfully achieved ongoing pregnancies with the oocytes retained for reproductive purposes. Mature oocytes, obtained within 2 hours following transvaginal aspiration, were enucleated using one of two methods, extrusion or aspiration, after 45 minutes of incubation in cytochalasin B. Rates of oocyte lysis or degeneration did not differ between the two methods. Somatic cell nuclear transfer (SCNT) embryos were constructed using two established adult male fibroblast lines of normal karyotype. High rates of pronuclear formation (66%), early cleavage (47%), and blastocyst (23%) development were observed following incubation in standard in vitro fertilization culture media. One cloned blastocyst was confirmed by DNA and mitochondrial DNA fingerprinting analyses, and DNA fingerprinting of two other cloned blastocysts indicated that they were also generated by SCNT. Blastocysts were also obtained from a limited number of parthenogenetically activated oocytes. This study demonstrates, for the first time, that SCNT can produce human blastocyst-stage embryos using nuclei obtained from differentiated adult cells and provides new information on methods that may be needed for a higher level of efficiency for human nuclear transfer. STEM CELLS 2008;26:485-493 Disclosure of potential conflicts of interest is found at the end of this article.

SummaryReasons for performing studyLungeing is commonly used as part of standard lameness examinations in horses. Knowledge of how lungeing influences motion symmetry in sound horses is needed.ObjectivesThe aim of this study was to objectively evaluate the symmetry of vertical head and pelvic motion during lungeing in a large number of horses with symmetric motion during straight line evaluation.Study designCross‐sectional prospective study.MethodsA pool of 201 riding horses, all functioning well and considered sound by their owners, were evaluated in trot on a straight line and during lungeing to the left and right. From this pool, horses with symmetric vertical head and pelvic movement during the straight line trot (n = 94) were retained for analysis. Vertical head and pelvic movements were measured with body mounted uniaxial accelerometers. Differences between vertical maximum and minimum head (HDmax, HDmin) and pelvic (PDmax, PDmin) heights between left and right forelimb and hindlimb stances were compared between straight line trot and lungeing in either direction.ResultsVertical head and pelvic movements during lungeing were more asymmetric than during trot on a straight line. Common asymmetric patterns seen in the head were more upward movement during push‐off of the outside forelimb and less downward movement during impact of the inside limb. Common asymmetric patterns seen in the pelvis were less upward movement during push‐off of the outside hindlimb and less downward movement of the pelvis during impact of the inside hindlimb. Asymmetric patterns in one lunge direction were frequently not the same as in the opposite direction.ConclusionsLungeing induces systematic asymmetries in vertical head and pelvic motion patterns in horses that may not be the same in both directions. These asymmetries may mask or mimic fore‐ or hindlimb lameness.