Chemopreventive effect of a milk whey by-product derived from Buffalo (Bubalus bubalis) in protecting from colorectal carcinogenesis.

BACKGROUND: Several studies show that natural foods are a source of compounds with anticancer properties that affect the gut microbiota and its metabolites. In the present study, we investigate the effect of a delactosed buffalo milk whey by-product (DMW) on colorectal carcinogenesis. METHODS: The effect of DMW on colorectal carcinoma (CRC) was investigated in the established mouse model of azoxymethane (AOM)-induced colon carcinoma, which closely resembles the human clinical condition of CRC. The effect of DMW on CRC immortalized cell lines was also evaluated to further identify the antineoplastic mechanism of action. RESULTS: Pretreatment of AOM-treated mice with DMW significantly (P < 0.05) reduced the percentage of mice bearing both aberrant crypt foci with more than four crypts (which are early precancerous lesions that progress to CRC) and tumors. In addition, DMW completely counteracted the effect of AOM on protein expression of caspase-9, cleaved caspase-3 and poly ADP-ribose polymerase in colonic tissue. Administration of DMW alone (i.e. without AOM) resulted in changes in the composition of the gut microbiota, leading to enrichment or depletion of genera associated with health and disease, respectively. DMW was also able to restore AOM-induced changes in specific genera of the gut microbiota. Specifically, DMW reduced the genera Atopobiaceae, Ruminococcus 1 and Lachnospiraceae XPB1014 and increased the genera Parabacteroides and Candidatus Saccharimonas, which were increased and reduced, respectively, by AOM. Blood levels of butyric acid and cancer diagnostic markers (5-methylcytidine and glycerophosphocholine), which were increased by AOM treatment, were reduced by DMW. Furthermore, DMW exerted cytotoxic effects on two human CRC cell lines (HCT116 and HT29) and these effects were associated with the induction of apoptotic signaling. CONCLUSIONS: Our results suggest that DMW exerts chemopreventive effects and restores the gut microbiota in AOM-induced CRC, and induces cytotoxic effect on CRC cells. DMW could be an important dietary supplement to support a healthy gut microbiota and reduce the prevalence of CRC in humans. Video Abstract.

Lack of effect of melatonin on ovarian function and response to estrous synchronization and fixed-time AI during the nonbreeding season in lactating dairy buffalo (Bubalus bubalis).

The present study was conducted to examine whether pretreatment with melatonin would enhance ovarian follicular functions and increase response to estrous synchronization and fixed-time AI (TAI) during the nonbreeding season in lactating dairy buffalo. In Experiment 1, buffalo cows without a detectable corpus luteum (CL) were assigned on Day -20 (D-20) to three groups: control (n = 12); melatonin (n = 13); progesterone (P4) (n = 15). Cows in the melatonin group were implanted with melatonin on D-20. From D0 to D9, there was imposing of an estrous synchronization treatment regimen using either a standard Ovsynch protocol (control, melatonin) or a P4-based Ovsynch treatment regimen (P4). There were no differences (P > 0.05) among groups for the presence of a CL at D0, size of the largest follicle at D0, ovulation to GnRH injection at D0 and D9, or the time to ovulation after injection of GnRH at D9. In Experiment 2, there was imposing of the same treatment regimens as in Experiment 1, with inclusion of TAI. Females of the P4 group had a greater (P = 0.001) pregnancy/AI percentage (60 %) than those in the control (17 %) and melatonin (23 %) groups. Females of the P4 group also had a larger (P = 0.005) CL at D20 compared with those in the control and melatonin groups. Findings indicate treatment with melatonin for 20 days did not affect ovarian functions or the response to an estrous synchronization treatment regimen and TAI during the nonbreeding season in lactating dairy buffalo.

Reproductive management in buffalo by artificial insemination.

Artificial insemination (AI) is important for genetic improvement and to control the period of breeding in buffalo and has increased significantly over the past 20 years. AI is more difficult in buffalo compared with cattle due to variable estrous cycles, reduced estrous behavior, and reproductive seasonality. The latter is associated with a higher incidence of anestrus and increased embryonic mortality during the nonbreeding season. Protocols to control the stage of the estrous cycle have undergone recent development in buffalo. These protocols are based on the control of both the luteal phase of the cycle, mainly by prostaglandins and progesterone, and follicle development and ovulation, by prostaglandins, progesterone, GnRH, hCG, eCG and estradiol. Protocols that synchronize the time of ovulation enable fixed timed AI, avoiding estrous detection. Factors to consider when selecting an AI protocol include animal category (heifers, primiparous or pluriparous), reproductive status (cyclic or anestrus), and season. This review looks at the current status of estrus synchronization and AI in buffalo and provides some practical suggestions for application of AI in the field.

Administration of PGF2α during the periovulatory period increased fertilization rate in superovulated buffaloes.

The aim of the present study was to determine the recovery of embryonic structures (ova/embryos) and fertilization rate in superovulated buffaloes treated with PGF2α during the periovulatory period. On day 0 (D0), buffaloes at random stages of the estrous cycle were treated with an intravaginal progesterone device (P4; 1.0 g) and estradiol benzoate (EB, 2.0 mg i.m.). From D4 to D7, all buffaloes received i.m. FSH (200 mg total) twice-daily over 4 days in decreasing doses. On D6 and D7, the animals were given PGF2α analogue (0.53 mg i.m. sodium cloprostenol) and the P4 device was removed on D7. On D8, all buffaloes received GnRH (20 µg i.m. buserelin acetate). Buffaloes were then randomly allocated to one of three groups: control (Group C, n = 18), no further treatment; PGF2α analogue injection (Group IM-PGF; n = 18), four injections (0.53 mg i.m. sodium cloprostenol) 12 h apart, from D8 to D10; PGF2α analogue osmotic pump (Group OP-PGF; n = 18), s.c. osmotic mini-pump (2.12 mg sodium cloprostenol) from D8 to D10. The study had a crossover design (three treatments x three replicates). All animals underwent timed AI, 12 and 24 h after treatment with GnRH. Embryonic structures were recovered on D14. Ovarian ultrasonography was used on D8 and D14 to record follicular superstimulation and superovulatory responses. Blood samples were obtained on Days 7, 8, 9 and 10 to measure circulating concentrations of P4, E2 and PGFM. Data were analyzed by GLIMMIX procedure of SAS®. There was no effect (P = 0.58) of treatment on the total number of embryonic structures (Group C, 2.1 ±â€¯0.8; Group IM-PGF, 2.1 ±â€¯0.6; Group OP-PGF, 1.4 ±â€¯0.4). There was also no effect (P = 0.93) of treatment on the recovery rate of embryonic structures (oocytes and embryos D14/CL D14). The fertilization rate was higher (P = 0.04) in Groups IM-PGF (84.6%) and OP-PGF (88.0%), which did not differ, than Group C (63.2%). The viable embryos rate was greater (P < 0.01) for Groups IM-PGF (82.0%) and OP-PGF (88.0%) than Group C (52.6%). There was no interaction between treatment and time and treatment effects for P4, E2 and PGFM concentrations. The findings showed that treatment with PGF2α during the periovulatory period has potential to increase fertilization rate and embryo production in superovulated buffaloes.

Global transcriptome profiles of Italian Mediterranean buffalo embryos with normal and retarded growth.

The transcriptome profiles were compared for buffalo embryos with normal growth and embryos with retarded growth on Day 25 after mating. Embryos with retarded growth on Day 25 after mating have a reduced likelihood of undergoing attachment to the uterine endometrium and establishing a pregnancy. Italian Mediterranean buffaloes were mated by AI and on Day 25 underwent trans-rectal ultrasonography to ascertain embryo development. Embryos with an embryonic width (EW)>2.7 mm were classed as normal embryos and embryos with an EW<2.7 mm were classed as retarded embryos. Three buffaloes with embryos of the largest EW (3.7, 3.7 and 3.9 mm) and three buffaloes with embryos of the smallest EW (1.5, 1.6 and 1.9 mm) were slaughtered on Day 27 to recover embryos for transcriptome analysis using a bovine custom designed oligo array. A total of 1,047 transcripts were differentially expressed between embryos with normal growth and embryos with retarded growth. Retarded embryos showed 773/1,047 (74%) transcripts that were down-regulated and 274/1,047 (26%) transcripts that were up-regulated relative to normal embryos; in silico analyses focused on 680/1,047 (65%) of the differentially expressed transcripts. The most altered transcripts observed in retarded embryos were associated with membrane structure and function and with metabolic and homeostasis maintenance functions. Other notable functions altered in retarded embryos were developmental processes and in particular nervous system differentiation and function. Specific biochemical pathways such as the complement cascade and coagulation were also altered in retarded embryos. It was concluded from the findings that buffalo embryos with retarded growth on Day 25 after mating show altered gene expression compared with normal embryos, and some de-regulated functions are associated with attachment to the uterine endometrium.