scholarly journals Molecular cues to the anti-implantation effect of nano-puerarin in rats

Reproduction ◽  
2016 ◽  
Vol 151 (6) ◽  
pp. 693-707 ◽  
Author(s):  
Ghungroo Saraswat ◽  
Rajdeep Guha ◽  
Kalyani Mondal ◽  
Piyali Saha ◽  
Sayani Banerjee ◽  
...  
Keyword(s):  
Oestrogen Receptor ◽  
Stromal Cell ◽  
Cell Activation ◽  
Drug Loading ◽  
Cyclin D3 ◽  
Receptor Subtypes ◽  
Decidual Cells ◽  
Dose Dependent

AbstractPuerarin, a selective oestrogen receptor modulator, intercepts implantation in rats, albeit at unacceptably higher doses. We developed poly lactic-co-glycolic acid-encapsulated nano-puerarin (PN) and mapped the molecular pathway underlying its anti-implantation effects. Smooth-surfaced and spherical PN having a mean diameter of ∼147nm was obtained with good yield, efficient encapsulation, and optimum drug loading. In culture, PN slowly and steadily released puerarin, which was readily taken up by the decidual cells. PN exerted a dose-dependent anti-implantation effect. As marked by attenuated expression of stromal cell desmin, alkaline phosphatase, IGFBP1, and decidual prolactin-related protein, the anti-implantation effect of PN seemed secondary to compromised decidualization. Usingin vivo(pregnant and pseudopregnant rats) andin vitro(endometrial stromal cell culture) treatment models, we document that PN enforced inhibition of uterine expression ofHbegfandHoxa10and their downstream signalling molecules, Cyclin D3 (CCND3)/CDK4. PN also efficiently ablated theIhh-Nr2f2-Bmp2signalling pathway and invited the loss of uterine potential for decidualization. There was a dose-dependent up-regulation of RHOA and its effector protein kinase, ROCK1, leading to the promotion of MLC phosphorylation and actin–myosin interaction. PN also down-regulated the stromal cell activation of ERK½ and expression of MMP9. These effects acting together stabilized the stroma and inhibited the stromal cell migration. Central to this array of events was the adversely altered endometrial expression of oestrogen receptor subtypes and repression of progesterone receptor that indulged endless proliferation of luminal epithelia and distorted the precisely choreographed stroma–epithelia crosstalk. Thus, PN dismantles the endometrial bed preparation and prevents implantation.

CATECHOL OESTROGENS AND GONADOTROPHIN SECRETION IN THE EWE: AFFINITY FOR PITUITARY OESTROGEN RECEPTORS IN VITRO AND ACTION ON GONADOTROPHIN SECRETION IN VIVO

1980 ◽  
Vol 85 (3) ◽  
pp. 503-509 ◽  
Author(s):  
I. J. CLARKE ◽  
J. K. FINDLAY
Keyword(s):  
Negative Feedback ◽  
Oestrogen Receptor ◽  
Dose Response Relationship ◽  
Concurrent Administration ◽  
Feedback Effects ◽  
Dependent Relationship ◽  
Gonadotrophin Secretion ◽  
Dose Dependent

The binding of three catechol oestrogens, 2-OH-oestradiol-17β, 4-OH-oestrone and 2-OH-oestrone, to the ovine pituitary oestrogen receptor was measured in vitro to establish doses for the assessment of the effects of catechol oestrogens in vivo. Relative to oestradiol (100%) the compounds had receptor affinities of 30, 20 and 5% respectively. A dose of oestradiol sufficient to cause negative-feedback effects on the secretion of LH and FSH in ovariectomized ewes was established by intracarotid (i.c.) injections of 0·625–5·0 μg/dose (n = 3), and by measuring plasma levels of gonadotrophins in jugular venous samples taken at intervals of 20 min from 3 h before until 4 h after injection. A dose-dependent relationship (r = 0·88, P<0·001) was found for oestradiol and plasma LH levels. Plasma FSH was slightly (12–25%) but significantly (P<0·05) reduced by doses of 1·25–5·0 μg oestradiol, but no dose–response relationship was observed. Ovariectomized ewes (n = 4/group) were given 2·5 μg oestradiol (i.c.) simultaneously with 83 μg 2-OH-oestradiol, 125 μg 4-OH-oestrone or 500 μg 2-OH-oestrone. These doses of catechol oestrogens were chosen as being ten times that of oestradiol, with the relative affinities for oestrogen receptor taken into account. Concurrent administration of such doses of catechol oestrogens had no effect on the negative-feedback action of oestradiol in vivo. We have concluded that catechol oestrogens in the circulation probably do not modulate the action of oestradiol on release of LH or FSH; this does not preclude a possible role for them as locally produced regulators of oestrogen action.

scholarly journals CD3-Activating Bi-Specific Antibody Targeting CD19 on B Cells in Mono- and Bi-Valent Format

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 4169-4169 ◽  
Author(s):  
Yumin CUI ◽  
Zhihua Huang ◽  
Xinfeng Zhang ◽  
Wuzhong Shen ◽  
Hanyang Chen ◽  
...  
Keyword(s):  
T Cell ◽  
B Cell ◽  
T Cell Activation ◽  
Specific Antibody ◽  
Cell Activation ◽  
Cell Killing ◽  
Cell Binding ◽  
Dose Dependent

Abstract Immunotherapies targeting B-lineage-specific surface marker CD19 had demonstrated promising clinical results. Two CD19 CAR-T therapies (Kymriah® and Yescarta®) have been approved by FDA to treat patients with B cell malignancies, however, the complicated manufacturing process and low throughput limit its accessibility to more patients, especially in developing countries. The first CD3-activating bi-specific antibody targeting CD19, Blincyto, or CD19 BiTE, was approved to treat relapsed and refractory acute lymphoblastic lymphoma (r/r ALL). The relatively short half-life of Blincyto requires continuous IV infusion for weeks to maintain a steady levels of drug exposure, not to mention the high risk of developing severe cytokine release syndrome in patients. We had established a bispecific antibody platform ITabTM (immunotherapy antibody) for the generation of CD3-activating bi-specific antibodies that could potentially overcome the shortcomings of BiTEs. A CH1 domain was introduced into the ITabTM construct design with the intent to increase the molecular weight thus led to extend the serum half-life of the bispecific antibody. A novel CD3-activating and monkey cross-reactive antibody was generated with a less degree of T cell activation and cytokine release compared to BiTEs. A bi-valent binding to tumor associated antigen (TAA) format was established to target tumor cells and/or stem cells expressing very low levels of TAA. We report here the biological properties of the mono-valent/bi-valent binding of CD19 bi-specific antibody with CD3-activating activity (A-319/A-329). A series of studies were conducted to evaluate the bioactivities of A-319/A-329 in vitro and in vivo including binding to CD3 and CD19 antigens, T-cell and B-cell binding activities, T cell activation and proliferation and B cell killing activities in vitro as well as in vivo efficacy using human PBMC engrafted mouse xenograft models. The in vitro data showed that the mono-valent and bi-valent CD19 binding had little effect on the CD3-associated activities including CD3 antigen binding affinity, T cell binding and T cell activation. In contrast, the bi-valent binding format A-329 showed better potency compared to the mono-valent format A-319 in CD19 binding (KD 0.89 nM vs 19.4 nM); B cell binding (EC50 at 2.3 pM vs 462 pM); in vitro human B cell killing (EC50 0.2 pM vs 3.4 pM). Both A-319 and A-329 were capable of mediating tumor cell lysis with EC50 at 0.03~4 pM. A-329 demonstrated a greater killing activity on Pfeiffer, a human diffuse large B-cell lymphoma (DLBCL) cell line with a low expression of CD19 antigen. In human PBMC engrafted NOG mouse xenograft model, a dose-dependent tumor growth inhibition was observed at 0.5~100 µg/kg in both A-319 and A-329. In monkey studies, when A-319 and A-329 was dosed at 3, 10, 30 µg/kg, twice or three times weekly via IV infusion for A-329 or A-319. Dose-dependent elimination of peripheral blood B cells were observed with both ITabTM. The CD19 bi-valent format of A-329 revealed more complete B cell killing in monkeys. No significant difference of cytokine induction or liver injuries were observed between A-319 and A-329. These results demonstrated that both A-319 and A-329 may benefit patients with B cell malignancies with less dosing frequency and lower cytokine inductions especially, A-329 may have the potential to targeting the low CD19 expressing tumor stem cells. Disclosures No relevant conflicts of interest to declare.

scholarly journals Overexpression of Cyclin D3 Improves Decidualization Defects in Hoxa-10−/− Mice

Endocrinology ◽  
2012 ◽  
Vol 153 (11) ◽  
pp. 5575-5586 ◽  
Author(s):  
Julie M. Sroga ◽  
Fei Gao ◽  
Xinghong Ma ◽  
Sanjoy K. Das
Keyword(s):  
In Situ Hybridization ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Regulatory Protein ◽  
Cyclin D3 ◽  
In Vivo Studies ◽  
Phase Cell

Abstract Uterine decidualization, a crucial process for implantation, is a tightly regulated process encompassing proliferation, differentiation, and polyploidization of uterine stromal cells. Hoxa (Homeobox A)-10, a homeobox transcription factor, is highly expressed in decidualizing stromal cells. Targeted gene deletion experiments have demonstrated marked infertility resulting from severely compromised decidualization in Hoxa-10−/− mice. However, the underlying mechanism by which Hoxa-10 regulates stromal cell differentiation remains poorly understood. Cyclin D3, a G1 phase cell-cycle regulatory protein involved in stromal cell proliferation and decidualization, is significantly reduced in Hoxa-10−/− mice. The expression of cyclin D3 in the pregnant mouse uterus parallels stromal cell decidualization. Here, we show that adenovirus-driven cyclin D3 replacement in Hoxa-10−/− mice improves stromal cell decidualization. To address our question of whether cyclin D3 replacement in Hoxa-10−/− mice can improve decidualization, both in vitro and in vivo studies were completed after the addition of cyclin D3 or empty (control) viral vectors. Immunostaining demonstrated increased proliferation and decidualization in both in vitro and in vivo studies, and in situ hybridization confirmed increased expression of decidualization markers in vivo. Placentation was demonstrated as well in vivo in the cyclin D3-replaced animals. However, fertility was not restored in Hoxa-10−/− mice after d 10 of pregnancy. Finally, we identified several downstream targets of cyclin D3 during decidualization in vitro via proteomics experiments, and these were confirmed using in situ hybridization in vivo. Collectively, these results demonstrate that cyclin D3 expression influences a host of genes involved in decidualization and can improve decidualization in Hoxa-10−/− mice.

Immunosuppression, nonspecific B-cell activation, and mitogenic activity associated with a high molecular weight component from Listeria monocytogenes

1982 ◽  
Vol 28 (12) ◽  
pp. 1373-1381 ◽  
Author(s):  
T. V. Otokunefor ◽  
S. B. Galsworthy
Keyword(s):  
Molecular Weight ◽  
Listeria Monocytogenes ◽  
High Molecular Weight ◽  
Cell Activation ◽  
B Cell Activation ◽  
Saline Extract ◽  
High Molecular Weight Component ◽  
Dose Dependent

A high molecular weight component of a saline extract derived from Listeria monocytogenes contained amino acids, carbohydrates, and phosphorus. The same fraction was capable of promoting both the in vitro mitogenic and adjuvant activities and the in vivo immunosuppressive activity displayed by the crude extract. The material was mitogenic to B but not to T lymphocytes in vitro. Responses to sheep and horse erythrocytes as well as to lipopolysaccharide were suppressed. Immunosuppression was dose dependent and was present at 1, 2, or 3 days but absent 7 days after injection. Both primary and secondary responses to sheep erythrocytes were impaired.

scholarly journals 863 In vitro and in vivo studies establish DuoBody®-CD3xB7H4 as a novel drug candidate for the treatment of solid cancers

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A904-A904
Author(s):  
Louise Koopman ◽  
Laura Smits-de Vries ◽  
Frederikke Lihme Egerod ◽  
Sebastiaan Wubben ◽  
Mischa Houtkamp ◽  
...  
Keyword(s):  
T Cell ◽  
Antitumor Activity ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cynomolgus Monkeys ◽  
Animal Care ◽  
Dose Dependent

BackgroundThe immune checkpoint protein B7H4 is expressed on malignant cells in various solid cancers, whereas its expression is highly restricted in normal tissue. B7H4 is therefore an attractive target for a CD3 bispecific antibody (bsAb) therapeutic. Moreover, its expression is reported to be inversely correlated with PD-L1. Here, we describe the preclinical characterization of two B7H4-targeting CD3 bsAbs with different CD3 affinities, supporting the selection of our clinical lead, DuoBody-CD3xB7H4 (GEN1047).MethodsB7H4 protein expression in patient-derived samples was determined by immunohistochemistry. Controlled Fab-arm exchange of an Fc-silenced B7H4 antibody with two Fc-silenced CD3ε-binding antibodies generated two CD3xB7H4 bsAbs that differ in CD3 binding affinity by approximately 30-fold. In vitro T-cell mediated cytotoxicity, T-cell activation, and cytokine release were assayed using cocultures of B7H4-expressing tumor cells and healthy donor T cells. Nonclinical safety (NCS) of the two CD3xB7H4 bsAbs was assessed in cynomolgus monkeys, and antitumor activity of the clinical lead in vivo was tested in a patient-derived xenograft (PDX) screen in mice with a humanized immune system (HIS).ResultsB7H4 protein expression was confirmed in tumor biopsies from multiple indications, including breast, ovarian and lung cancer. Both bsAbs induced target-specific and dose-dependent tumor cell kill in vitro. Maximal kill and T-cell activation were comparable for both variants, although the potency of the high CD3 affinity bsAb was higher. However, production of inflammatory cytokines at comparable effective concentrations (IC90) was lower for the low CD3 affinity bsAb. Single dose NCS studies in cynomolgus monkeys showed that both CD3xB7H4 bsAbs were well-tolerated. A dose-dependent increase in plasma cytokines IL-6 and MCP-1 2 hours after dosing was observed only with the high CD3 affinity bsAb. Based on these findings, the low CD3 affinity bsAb was selected for follow-up studies and named DuoBody-CD3xB7H4 (GEN1047). DuoBody-CD3xB7H4 demonstrated antitumor activity in vivo in a PDX screen in HIS mice. Repeated dosing of DuoBody-CD3xB7H4 in cynomolgus monkeys confirmed an acceptable safety profile up to the maximal dose tested (30 mg/kg).ConclusionsThese studies describe the preclinical development of DuoBody-CD3xB7H4, a bsAb that induces T-cell mediated cytotoxicity of B7H4-positive tumor cells, which may provide an alternative therapeutic modality in the immune-oncology space for patients with solid cancers.Ethics ApprovalAnimal experiments were performed according to the guidelines of the Institutional Animal Care and Use Committee (IACUC) and in accordance with the regulations of the Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC). NCS studies were conducted at Citoxlab (Evreux, France) and Charles River Laboratories (Tranent, UK) in accordance with the European Convention for the Protection of Vertebrate Animals Used for Experimental and Other Scientific Purposes (Council of Europe).

Soluble Thrombomodulin Purified from Human Urine Exhibits a Potent Anticoagulant Effect In Vitro and In Vivo

1995 ◽  
Vol 73 (05) ◽  
pp. 805-811 ◽  
Author(s):  
Yasuo Takahashi ◽  
Yoshitaka Hosaka ◽  
Hiromi Niina ◽  
Katsuaki Nagasawa ◽  
Masaaki Naotsuka ◽  
...  
Keyword(s):  
Human Urine ◽  
Specific Activity ◽  
Anticoagulant Activity ◽  
Rabbit Lung ◽  
Soluble Thrombomodulin ◽  
Molecular Weights ◽  
Dose Dependent Fashion ◽  
Dose Dependent

SummaryWe examined the anticoagulant activity of two major molecules of soluble thrombomodulin purified from human urine. The apparent molecular weights of these urinary thrombomodulins (UTMs) were 72,000 and 79,000, respectively. Both UTMs showed more potent cofactor activity for protein C activation [specific activity >5,000 thrombomodulin units (TMU)/mg] than human placental thrombomodulin (2,180 TMU/mg) and rabbit lung thrombomodulin (1,980 TMU/mg). The UTMs prolonged thrombin-induced fibrinogen clotting time (>1 TMU/ml), APTT (>5 TMU/ml), TT (>5 TMU/ml) and PT (>40 TMU/ml) in a dose-dependent fashion. These effects appeared in the concentration range of soluble thrombomodulins present in human plasma and urine. In the rat DIC model induced by thromboplastin, administration of UTMs by infusion (300-3,000 TMU/kg) restored the hematological abnormalities derived from DIC in a dose-dependent fashion. These results demonstrate that UTMs exhibit potent anticoagulant and antithrombotic activities, and could play a physiologically important role in microcirculation.

Vasorelaxant and Antihypertensive Effects of Mentha pulegium L. in Rats: An in vitro and in vivo Approach

2020 ◽  
Vol 20 ◽  
Author(s):  
Mohammed Ajebli ◽  
Mohamed Eddouks
Keyword(s):  
Blood Pressure ◽  
Acute Experiment ◽  
Antihypertensive Activity ◽  
Mentha Pulegium ◽  
In Vitro Experiment ◽  
Arterial Blood ◽  
Dose Dependent ◽  
Ca2 Channels

Aims and objective: The aim of the study was to investigate the effect of aqueous aerial part extract of Mentha pulegium L. (Pennyrile) (MPAE) on arterial pressure parameters in rats. Background: Mentha pulegium is a medicinal plant used to treat hypertension in Morocco. Material and methods: In the current study, MPAE was prepared and its antihypertensive activity was pharmacologically investigated. L-NAME-hypertensive and normotensive rats have received orally MPAE (180 and 300 mg/kg) during six hours for the acute experiment and during seven days for the sub-chronic treatment. Thereafter, systolic, diastolic, mean arterial blood pressure and heart rate were evaluated. While, in the in vitro experiment, isolated denuded and intact thoracic aortic rings were suspended in a tissue bath system and the tension changes were recorded. Results: A fall in blood pressure was observed in L-NAME-induced hypertensive treated with MPAE. The extract also produced a dose-dependent relaxation of aorta pre-contracted with NE and KCl. The study showed that the vasorelaxant ability of MPAE seems to be exerted through the blockage of extracellular Ca2+ entry. Conclusion: The results demonstrate that the extract of pennyrile exhibits antihypertensive activity. In addition, the effect may be, at least in part, due to dilation of blood vessels via blockage of Ca2+ channels.

Pharmacology of 5-HT1A Receptors: Critical Aspects

CNS Spectrums ◽  
1998 ◽  
Vol 3 (10) ◽  
pp. 17-38 ◽  
Author(s):  
Franco Borsini
Keyword(s):  
Receptor Subtypes ◽  
Laboratory Conditions ◽  
In Vivo Effects ◽  
Critical Aspects

AbstractMyriad difficulties exist in analyzing the pharmacology of the serotonin 1A (5-HT1A) receptor. The receptor may demonstrate a different activity depending on the tissue or species used for analysis, the agent used, laboratory conditions, and differences between in vitro and in vivo effects of compounds. Affinity for 5-HT receptors also varies widely, presenting difficulties in drawing definitive conclusions on affinity values for various compounds. At least two possibilities exist to explain the diversity of pharmacology of 5-HT receptors. First, it is possible that different 5-HT1A receptor subtypes exist. Second, the 5-HT1A receptors may play a far more complex role than previously believed.

scholarly journals Long-Acting Risperidone Dual Control System: Preparation, Characterization and Evaluation In Vitro and In Vivo

Pharmaceutics ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1210
Author(s):  
Xieguo Yan ◽  
Shiqiang Wang ◽  
Kaoxiang Sun
Keyword(s):  
Drug Loading ◽  
Entrapment Efficiency ◽  
In Vitro Dissolution ◽  
Dissolution Behavior ◽  
In Vivo Evaluation ◽  
Long Term Treatment ◽  
Long Acting

Schizophrenia, a psychiatric disorder, requires long-term treatment; however, large fluctuations in blood drug concentration increase the risk of adverse reactions. We prepared a long-term risperidone (RIS) implantation system that can stabilize RIS release and established in-vitro and in-vivo evaluation systems. Cumulative release, drug loading, and entrapment efficiency were used as evaluation indicators to evaluate the effects of different pore formers, polymer ratios, porogen concentrations, and oil–water ratios on a RIS implant (RIS-IM). We also built a mathematical model to identify the optimized formulation by stepwise regression. We also assessed the crystalline changes, residual solvents, solubility and stability after sterilization, in-vivo polymer degradation, pharmacokinetics, and tissue inflammation in the case of the optimized formulation. The surface of the optimized RIS microspheres was small and hollow with 134.4 ± 3.5 µm particle size, 1.60 SPAN, 46.7% ± 2.3% implant drug loading, and 93.4% entrapment efficiency. The in-vitro dissolution behavior of RIS-IM had zero-order kinetics and stable blood concentration; no lag time was released for over three months. Furthermore, the RIS-IM was not only non-irritating to tissues but also had good biocompatibility and product stability. Long-acting RIS-IMs with microspheres and film coatings can provide a new avenue for treating schizophrenia.

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