Production of CaCO3-single-coated probiotics and evaluation of their spectroscopic properties, morphological characteristics, viability, and intestinal delivery efficiency.

The intake of probiotics offers various health benefits; however, their efficacy depends on the maintenance of viability during industrial processing and digestion. Probiotic viability can be compromised during encapsulation, freeze-drying, storage, and digestion, necessitating multiple coatings. This complicates production and raises costs. In this study, CaCO3-single-coated probiotics (CSCPs) were prepared, an approach rarely reported before. Through instrumental analyses, the encapsulation of probiotics within CaCO3 was confirmed, ensuring their high viability. This proposed technology effectively preserves the viability of probiotics during the encapsulation and freeze-drying processes, resulting in minimal cell loss. Moreover, CSCPs demonstrated exceptional viability performance under simulated gastric and intestinal conditions. Notably, 100% of these microorganisms reached the intestines, delivering over 10 billion CFUs of probiotics in a viable state. This study highlights the potential of CSCPs as a feasible solution for overcoming probiotic encapsulation challenges and optimizing therapeutic benefits.

Immunostimulatory activity and safety evaluation of Bordetella bronchiseptica-derived lipopolysaccharide, a new vaccine adjuvant candidate.

Adjuvants are used to elicit strong immune responses for vaccines that show poor immunogenicity. Previously, we demonstrated that a sonicated bacterin of Bordetella bronchiseptica can be used as a safe adjuvant that enhances the antigen-presenting capability of dendritic cells (DCs). In this study, we purified the lipopolysaccharide (LPS) of B. bronchiseptica (Bb-LPS) and investigated its immunogenic effects on DCs compared to those of Escherichia coli O26:B6 (O26)-derived LPS (O26-LPS), a positive control. Bb-LPS was purified using an LPS extraction kit. Limulus amebocyte lysate assay was performed to determine the optimal concentration of Bb-LPS and O26-LPS for treatment. Bb-LPS increased the metabolic activity of DCs at a concentration of 0 to 250 EU/mL, similar to that of O26-LPS. Bb-LPS significantly increased the expression level of CD40 and CD54, related to the immune responses of DCs. Bb-LPS enhanced the antigen-presenting capability of DCs and significantly increased the interferon-gamma/interleukin-4 ratio of CD4+ T cells co-cultured with DCs to 0.95 (p < 0.05). Moreover, Bb-LPS increased the production of pro-inflammatory cytokines in a safer manner than that obtained by O26-LPS. In vivo safety tests revealed that Bb-LPS was less toxic than O26-LPS in mice. This study demonstrated that Bb-LPS showed unique immune characteristics and immunogenic effects on the antigen-presenting capability of DCs, which differed from those of O26-LPS. This study provides valuable information for basic and clinical research for developing safe vaccine adjuvants.

Bordetella bronchiseptica is a potent and safe adjuvant that enhances the antigen-presenting capability of dendritic cells.

We previously demonstrated that Bordetella bronchiseptica (B. bronchiseptica) antigen (Ag) enhances the Mycoplasma hyopneumoniae Ag-specific immune response. The focus of this study was whether acellular bacterin of B. bronchiseptica could be used as an adjuvant to increase antigen-presenting capability of dendritic cells (DCs) by increasing the level of activation. The metabolic activity of DCs was increased by B. bronchiseptica, similar to lipopolysaccharide (LPS). Flow cytometry analysis revealed that B. bronchiseptica increases the expression of major histocompatibility complex class-2, cluster of differentiation (CD)40, CD54, and CD86 which are closely related to DC-mediated immune responses. B. bronchiseptica enhanced the production of cytokines related to adaptive immune responses. Furthermore, the survival rate of B. bronchiseptica-injected groups was 100% at 15 and 20 mg/kg doses, whereas that of LPS-injected groups was only 20%, 0% at 15 and 20 mg/kg doses respectively, and so B. bronchiseptica is likely to be safer than LPS. Taken together, these results indicate that B. bronchiseptica can be used as an adjuvant to enhance the antigen-presenting capability of DCs. B. bronchiseptica is a candidate for producing vaccines, especially in case of DC-mediating efficacy and safety demands. This study provides researchers and clinicians with valuable information regarding the usage of B. bronchiseptica as a safe bacteria-derived immunostimulating agent for developing efficient vaccines.

Subclinical hypothyroidism diagnosed by thyrotropin-releasing hormone stimulation test in infertile women with basal thyroid-stimulating hormone levels of 2.5 to 5.0 mIU/L.

OBJECTIVE: To investigate the prevalence of subclinical hypothyroidism (SH) diagnosed by thyrotropin-releasing hormone (TRH) stimulating test in infertile women with basal thyroid-stimulating hormone (TSH) levels of 2.5 to 5.0 mIU/L. METHODS: This study was performed in 39 infertile women with ovulatory disorders (group 1) and 27 infertile women with male infertility only (group 2, controls) who had basal serum TSH levels of 2.5 to 5.0 mIU/L and a TRH stimulating test. Serum TSH levels were measured before TRH injection (TSH0) and also measured at 20 minutes (TSH1) and 40 minutes (TSH2) following intravenous injection of 400 µg TRH. Exaggerated TSH response above 30 mIU/L following TRH injection was diagnosed as SH. Group 1 was composed of poor responders (subgroup A), patients with polycystic ovary syndrome (subgroup B) and patients with WHO group II anovulation except poor responder or polycystic ovary syndrome (subgroup C). RESULTS: The prevalence of SH was significantly higher in group 1 of 46.2% (18/39) compared with 7.4% (2/27) in group 2 (P=0.001). TSH0, TSH1, and TSH2 levels were significantly higher in group 1 than the corresponding values in group 2 (P<0.001, P<0.001, P<0.001). In group 1, TSH1 and TSH2 levels were significantly lower in subgroup C compared with those in subgroup A and B (P=0.008, P=0.006, respectively). CONCLUSION: TRH stimulation test had better be performed in infertile women with ovulatory disorders who have TSH levels between 2.5 and 5.0 mIU/L for early detection and appropriate treatment of SH.

The effect of luteal phase progesterone supplementation on natural frozen-thawed embryo transfer cycles.

OBJECTIVE: To evaluate the effect of progesterone supplementation during the luteal phase on pregnancy outcome in natural frozen-thawed embyo transfer (FTET) cycles. METHODS: In this retrospective cohort study, 228 consecutive patients who underwent FTET cycles between January 2009 and September 2012 were included. One hundred forty-five patients received luteal progesterone support (P group) but 83 patients did not receive any progesterone supplementation during luteal phase (control group). RESULTS: There were no differences in patients' characteristics between the two groups. The two groups were similar with respect to the characteristics of previous fresh in vitro fertilization cycle in which embryos were cryopreserved including the numbers of oocytes retrieved, mature oocytes, fertilized oocytes, grade 1 or 2 embryos and frozen embryos. Also, significant differences were not observed between the P and control groups in clinical pregnancy rate, embryo implantation rate and multiple pregnancy rate. However, miscarriage rate was significantly lower in the P group and live birth rate was significantly higher in the P group than in the control group (P<0.05, P<0.05). CONCLUSION: Our results suggest that luteal phase progesterone supplementation decreases miscarriage rate and improves live birth rate in natural FTET cycles.

Surgical resection or aspiration with ethanol sclerotherapy of endometrioma before in vitro fertilization in infertilie women with endometrioma.

OBJECTIVE: To evaluate whether the surgical resection or aspiration with ethanol sclerotherapy (AEST) of endometrioma before in vitro fertilization (IVF) affect controlled ovarian stimulation (COS) and IVF outcome in the infertilie women with endometroma undergoing IVF. METHODS: In this retrospective cohort study, 101 consecutive IVF/intracytoplasmic sperm injection cycles that were performed in 101 patients with endometrioma(s) between January 2008 and December 2012 were included. Before IVF, 36 patients underwent surgical resection of endometrioma (resection group), 29 patients had transvaginal endometrioma AEST (aspiration group), and 36 patients did not take any surgical intervention (control group). The three groups were compared in terms of COS and IVF outcomes. RESULTS: Total antral follicle count was significantly lower in the resection group than in the aspiration or control group. The numbers of follicles with a diameter of 14 to 17 mm on the human chorionic gonadotropin day, retrieved oocytes, mature oocytes, and fertilized oocytes were significantly lower in the resection group than in two other groups. However, three groups were similar in terms of clinical pregnancy rate (CPR) per initiated cycle, CPR per embryo transfer, embryo implantation rate, and miscarriage rate. CONCLUSION: Neither of surgical resection and AEST of endometrioma before IVF treatment can give any beneficial effect on IVF outcomes. Moreover, surgical resection of endometrioma can affect the ovarian reserve and ovarian response during COS.

The effect of aromatase inhibitor letrozole incorporated in gonadotrophin-releasing hormone antagonist multiple dose protocol in poor responders undergoing in vitro fertilization.

OBJECTIVE: To evaluate whether letrozole incorporated in a gonadotrophin-releasing hormone (GnRH) antagonist multiple dose protocol (MDP) improved controlled ovarian stimulation (COS) and in vitro fertilization (IVF) results in poor responders who underwent IVF treatment. METHODS: In this retrospective cohort study, a total of 103 consecutive IVF cycles that were performed during either the letrozole/GnRH antagonist MDP cycles (letrozole group, n=46) or the standard GnRH antagonist MDP cycles (control group, n=57) were included in 103 poor responders. COS results and IVF outcomes were compared between the two groups. RESULTS: Total dose and days of recombinant human follicle stimulating hormone (rhFSH) administered were significantly fewer in the letrozole group than in the control group. Duration of GnRH antagonist administered was also shorter in the letrozole group. The number of oocytes retrieved was significantly higher in the letrozole group. However, clinical pregnancy rate per cycle initiated, clinical pregnancy rate per embryo transfer, embryo implantation rate and miscarriage rate were similar in the two groups. CONCLUSION: The letrozole incorporated in GnRH antagonist MDP may be more effective because it results comparable pregnancy outcomes with shorter duration and smaller dose of rhFSH, when compared with the standard GnRH antagonist MDP.

Cathepsin B in eutopic and ectopic endometrial tissues of patients with endometriosis.

This study was performed to investigate the expression of cathepsin B mRNA and protein in eutopic and ectopic endometrial tissues of patients with endometriosis and in normal endometrial tissues and to clarify the association between the cathepsin B expression and endometriosis. A total of 40 women with histologically confirmed endometriosis were recruited for study group. For controls, 20 women undergoing operative treatment for uterine myoma, cervical intraepithelial neoplasia (CIN) or benign gynecologic conditions other than endometriosis were recruited. Eutopic endometrial tissues of both groups and ectopic endometrial tissue of study group were collected during the operations. We employed real time reverse transcriptase - polymerase chain reaction (RT-PCR) to quantify mRNA levels of cathepsin B in these tissues. Then, we performed western blot analysis to measure the protein levels of cathepsin B. The expressions of cathepsin B mRNA and protein were significantly higher in both eutopic and ectopic endometrial tissues of women with endometriosis than in endometrial tissues of controls. These data suggest that the higher expression of cathepsin B in the endometrial tissues might be associated with the development of endometriosis. In addition, eutopic endometrium itself with higher expression cathepsin B may play a pivotal role in the histogenesis of endometriosis.

Effect of Pioglitazone on Production of Regulated upon Activation Normal T-cell Expressed and Secreted (RANTES) and IVF Outcomes in Infertile Women with Endometriosis.

This study was performed to investigate the effect of peroxisome proliferators activated receptor-γ (PPAR-γ) ligand, pioglitazone, on production of regulated upon activation normal T-cell expressed and secreted (RANTES) and in vitro fertilization (IVF) outcome in infertile patients with endometriosis. Sixty-four infertile patients with stage III or IV endometriosis undergoing IVF were randomly allocated to the study or the control group. The long protocol of GnRH agonist (GnRH-a) was used for controlled ovarian stimulation (COS) in all patients. Patients in the study group were treated with pioglitazone at a dose of 15 mg/day orally from the starting day of GnRH-a treatment to the day of hCG injection. Blood samples were drawn for serologic assay of RANTES on the first day of GnRH-a treatment and the day of hCG injection. There were no differences between the study and control groups in patient characteristics. There were also no differences between the two groups in COS duration, and the numbers of retrieved oocytes, fertilized oocytes and embryos transferred. The clinical pregnancy rate per cycle was higher in the study group, but this difference was not statistically significant. However, embryo implantation rate was significantly higher in the study group of 12.5% compared with 8.6% in the control group (P<0.05). The serum RANTES levels after pioglitazone treatment were significantly lower than those before pioglitazone treatmen in the study group (P<0.05). Our data suggest that pioglitazone treatment can suppress RANTES production and improve the embryo implantation rate in patients with endometriosis undergoing IVF.

The Effect of Fibroblast Co-culture on In Vitro Maturation of Mouse Preantral Follicles.

This study was performed to evaluate the effects of fibroblast co-culture on in vitro maturation (IVM) of prepubertal mouse preantral follicles. The intact preantral follicles were obtained from the ovaries of 12-14 day old mice and these were cultured individually in α-minimal essential medium (α-MEM) supplemented with 5% fetal bovine serum (FBS), 100 mIU/Ml recombinant follicle stimulating hormone (rFSH), 1% insulin-transferrin-selenium, 100 µg/ml penicillin and 50 µg/Ml streptomycin as base medium for 12 days. A total of 200 follicles were cultured in base medium co-cultured with mouse embryonic fibroblast (MEF) (MEF group) (n=100) or only base medium as control group (n=100). Survival rate of follicles on day 12 of culture were significantly higher in the MEF group of 90.0%, compared with 77.0% of the control group (p=0.021). Follicle diameters on day 6 and 8 of the culture period were significantly larger in the MEF group than those in the control group (p=0.021, p=0.007, respectively). Estradiol levels in culture media on day 4, 6, 8, 10 and 12 of the culture period were significantly higher in the MEF group (p=0.043, p=0.021, p=0.006, p<0.001 and p=0.008, retrospectively). Our data suggest that MEF cell co-culture on IVM of mouse preantral follicle increases survival rate and promotes follicular growth and steroid production.