Journal: Biology of reproduction

Article Title: Conditional knockout of leptin receptor in the female reproductive tract reduces fertility due to parturition defects in mice.

doi: 10.1093/biolre/ioac062

Figure Lengend Snippet: Figure 1. Lepr cKO mice have reduced Lepr expression in the uterus and cervix compared with controls. (A) Real-time RT-PCR showed significant reductions in relative Lepr mRNA levels in the uterus (∗P < 0.0001) and cervix (∗P < 0.0001) of Lepr cKO mice (white bars) relative to controls (black bars). Bars represent steady-state mRNA levels relative to controls, and error bars represent ranges based on SEM of Ct values. (B-G) IHC for leptin receptor (brown) in (B) control uterus, (C) nonpregnant Lepr cKO uterus, (D) pregnant d6.5 Lepr cKO, L = lumen, G = gland, S = stroma. (E) Control ovary and (F) Lepr cKO ovary. (G) Negative control, with primary antibody omitted. F = follicle, CL = corpus luteum. (G) Negative control, no primary antibody. Scale bar in (B) for panels B–D,G. Scale bar in F for E–F.

Article Snippet: Briefly, sections were deparaffinized in a graded alcohol series, endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide solution, and nonspecific staining was blocked by using 1% horse serum, before overnight incubation at 4◦C with a goat anti-murine leptin receptor antibody, recognizing an epitope near the N-terminus present in all Lepr isoforms, at a 4 μg/mL in blocking buffer (Santa Cruz, SC-1835, RRID:AB_631883).

Techniques: Expressing, Quantitative RT-PCR, Control, Negative Control

Journal: Biology of reproduction

Article Title: Conditional knockout of leptin receptor in the female reproductive tract reduces fertility due to parturition defects in mice.

doi: 10.1093/biolre/ioac062

Figure Lengend Snippet: Figure 2. Fertility is impaired in Lepr cKO dams. (A) The number of pups born (live and dead) to Lepr cKO mothers was significantly lower than in controls over four parities (ANOVA, P = 0.0003). The number of pups per litter declined with increasing parity in dams of both genotypes (ANOVA, P = 0.02). (B) The total number of live pups born to Lepr cKO dams after four parities was significantly lower compared with controls (P = 0.002). (C) The number of days from pairing with a male to delivering a litter increased in Lepr cKO, but not control dams with increasing parity (ANOVA, genotype P = 0.005, parity P = 0.035, interaction P = 0.044). (D) The number of deliveries complicated by either dystocia or stillbirth in each of the four parities. In total the incidence of adverse advents was higher in Lepr cKO ∗P < 0.0001 (E) The total number of live and dead pups born across all four parities. ∗P < 0.0001 (F) Close up image of stillborn pup; pups appeared squished and flattened. In some instances, only pieces of pups were found, likely due to maternal cannibalism.

Article Snippet: Briefly, sections were deparaffinized in a graded alcohol series, endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide solution, and nonspecific staining was blocked by using 1% horse serum, before overnight incubation at 4◦C with a goat anti-murine leptin receptor antibody, recognizing an epitope near the N-terminus present in all Lepr isoforms, at a 4 μg/mL in blocking buffer (Santa Cruz, SC-1835, RRID:AB_631883).

Techniques: Control

Journal: Biology of reproduction

Article Title: Conditional knockout of leptin receptor in the female reproductive tract reduces fertility due to parturition defects in mice.

doi: 10.1093/biolre/ioac062

Figure Lengend Snippet: Figure 3. Estrous cycles and early pregnancy appear normal in Lepr cKO dams. (A) Average estrous cycle length in days. (B) Serum progesterone concentrations on pregnancy day 3.5. (C) The average number of implantation sites on pregnancy day 6.5. (D) Average CL numbers on pregnancy day 6.5.

Article Snippet: Briefly, sections were deparaffinized in a graded alcohol series, endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide solution, and nonspecific staining was blocked by using 1% horse serum, before overnight incubation at 4◦C with a goat anti-murine leptin receptor antibody, recognizing an epitope near the N-terminus present in all Lepr isoforms, at a 4 μg/mL in blocking buffer (Santa Cruz, SC-1835, RRID:AB_631883).

Techniques:

Journal: Biology of reproduction

Article Title: Conditional knockout of leptin receptor in the female reproductive tract reduces fertility due to parturition defects in mice.

doi: 10.1093/biolre/ioac062

Figure Lengend Snippet: Figure 4. Placental histology appears normal in Lepr cKO dams. Representative images of placentas at d17.5 from a (A) control and a (B) Lepr cKO dam. A full face, midsagittal section was stained with hematoxylin and eosin. No structural abnormalities, size differences, or signs of inflammation or hemorrhage were observed. The junctional zone is outlined in yellow (indicated by yellow letter J) and the labyrinth zone is outlined in black (indicated by black letter L).

Article Snippet: Briefly, sections were deparaffinized in a graded alcohol series, endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide solution, and nonspecific staining was blocked by using 1% horse serum, before overnight incubation at 4◦C with a goat anti-murine leptin receptor antibody, recognizing an epitope near the N-terminus present in all Lepr isoforms, at a 4 μg/mL in blocking buffer (Santa Cruz, SC-1835, RRID:AB_631883).

Techniques: Control, Staining

Journal: Biology of reproduction

Article Title: Conditional knockout of leptin receptor in the female reproductive tract reduces fertility due to parturition defects in mice.

doi: 10.1093/biolre/ioac062

Figure Lengend Snippet: Figure 5. Leptin receptor conditional knockout prolongs labor. (A) Average time from crouching to delivery of each pup. ∗P = 0.027. (B) Average time spent of labor per dam. † F test for variance, P < 0.01. (C) Average time from in labor per pup. †F test for variance, P = 0.03. (D) Proportion of pups stillborn (∗P = 0.04).

Article Snippet: Briefly, sections were deparaffinized in a graded alcohol series, endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide solution, and nonspecific staining was blocked by using 1% horse serum, before overnight incubation at 4◦C with a goat anti-murine leptin receptor antibody, recognizing an epitope near the N-terminus present in all Lepr isoforms, at a 4 μg/mL in blocking buffer (Santa Cruz, SC-1835, RRID:AB_631883).

Techniques: Knock-Out

Journal: Biology of reproduction

Article Title: Conditional knockout of leptin receptor in the female reproductive tract reduces fertility due to parturition defects in mice.

doi: 10.1093/biolre/ioac062

Figure Lengend Snippet: Figure 6. Cervical histology appears normal in Lepr cKO dams. Representative images of cervical sections immediately postpartum from control (A, C) and Lepr cKO (B, D) dams. Collagen matrix was visualized by Masson trichrome (A, B), and an image of an entire cervical longitudinal section was assembled. (C, D) PSR staining at a higher magnification. EC = ectocervix; EN = endocervix; I = location of inner images; O = location of outer images; S = stroma. Arrowheads point to epithelium.

Article Snippet: Briefly, sections were deparaffinized in a graded alcohol series, endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide solution, and nonspecific staining was blocked by using 1% horse serum, before overnight incubation at 4◦C with a goat anti-murine leptin receptor antibody, recognizing an epitope near the N-terminus present in all Lepr isoforms, at a 4 μg/mL in blocking buffer (Santa Cruz, SC-1835, RRID:AB_631883).

Techniques: Control, Staining

Journal: Frontiers in Pharmacology

Article Title: Biofunctionalization of cardiovascular stents to induce endothelialization: Implications for in- stent thrombosis in diabetes

doi: 10.3389/fphar.2022.982185

Figure Lengend Snippet: Summary of the recent studies investigating the use of biomolecules to induce stent endothelialization.

Article Snippet: , Stainless steel stents coated with murine monoclonal ENDs and CD34s (Beijing Lepu Medical Technology limited corporation, China), and SESs (Johnson & Johnson, United States). , Animal model: pigs. Findings: mean neointima area and ENDs, SESs and CD34s were lower in ENDs, SESs and CD34s when compared to DES at day 14 of implantation. Endothelial coverage was induced in ENDs and CD34 when compared to SESs and BMSs at days 7 and 14. , ( ) .

Techniques: Polymer, Combined Bisulfite Restriction Analysis Assay, In Vivo, In Vitro, Modification, Comparison, Animal Model, Clinical Proteomics, Introduce, Recombinant, Activity Assay, Activation Assay, Sonication, Incubation, Suspension, Migration, Genetically Modified, Sequencing, Solvent, Evaporation

Journal: BioMed Research International

Article Title: A Novel Stent Coated with Antibodies to Endoglin Inhibits Neointimal Formation of Porcine Coronary Arteries

doi: 10.1155/2014/428619

Figure Lengend Snippet: Histomorphometric analysis. Representative photos of haematoxylin-and-eosin-stained cross sections of arteries at 7 ((a), top panels) and 14 days ((b), top panels) after stent implantation (magnification, ×40). Bar graphs show mean neointima area ((a) and (b), bottom panels) and percent area stenosis (c) of stented arteries. Data represent the mean ± SEM ( n = 15). * P < 0.05 versus BMSs. BMSs: bare metal stents; SESs: sirolimus-eluting stents; and ENDs: endoglin antibody. Arrow indicated neointima.

Article Snippet: Thirty stainless steel stents coated with murine monoclonal anti-human endoglin antibody (ENDs) (Beijing Lepu Medical Technology limited corporation, China), thirty sirolimus-eluting stents (SESs) (purchased from Johnson & Johnson, USA), and thirty bare metal stents (BMSs) (purchased from Abbott, USA) were randomly assigned and placed in the left anterior descending, circumflex, or right coronary arteries (one stent per artery) of 30 pigs.

Techniques: Staining

Journal: Central European Journal of Immunology

Article Title: Experimental immunology Differences in the expression of leptin receptors on bone marrow and peripheral blood cells

doi: 10.5114/ceji.2012.32721

Figure Lengend Snippet: Fig. 1. Electronically differentiation according forward scat- ter (FS) vs. side scatter (SS) population of bone marrow cells (A) and peripheral blood leukocytes (B). Percentage of mononuclear cells (lymphocytes – gate A and monocytes – gate B) and granulocytes (gate C) are given in cytograms. Cytogram C shows granulocytes isolated from peripheral blood and stained with anti-CD13/CD33 (FL2) and mouse anti OB-R monoclonal antibodies. Goat antibody against mouse IgG con- jugated with fluoresceine (FL1) was used as the secondary anti- body. Area U1 contained CD13/33+Ob-R-cells; area U2 con- tained double positive cells: CD13/33+OB-R+ and in U3 area are cells double negative. In U4 area are another cells presented after isolation that are Ob-R+

Article Snippet: OB-R receptor was stained according to indirect immunofluorescence method using murine monoclonal IgG2A antibody against human leptin receptor manufactured by R&D Systems and secondary FITC-conjugated AffiniPure Goat Anti-Mouse IgG manufactured by Jackson ImmunoResearch Laboratories, Inc. Monocytic cells lineage was identified using anti CD4 and CD14 antibodies, while granulocytic lineage was identified with anti CD13 and CD33 antibodies.

Techniques: Isolation, Staining, Bioprocessing

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Leptin and soluble leptin receptor levels in obese and weight-losing individuals.

doi: 10.1210/jcem.87.4.8381

Figure Lengend Snippet: FIG. 1. Western blot analysis of sLR. Western blot analysis of plasma derived soluble leptin receptor, purified using anti-sLR Sepharose (lane 1), unreduced sLR-Fc (lane 2) and recombinant sLR-myc tagged (lane 3). In plasma, sLR was detected primarily as a band of approx- imately 180 kDa, and a very weak band at approximately 90–100 kDa. sLR-myc tagged was detected at approximately 90–100 kDa. Unreduced sLR-Fc showed a thick band at approximately 380 kDa. In this experiment, mAb 4C3 was used for detection. Similar results were obtained with mAb 2F1 (data not shown).

Article Snippet: Purification of native soluble leptin receptor from plasma To purify native sLR in a single step by affinity chromatography, the murine antihuman leptin receptor mAb 2F1 was coupled to CNBrSepharose beads (2 mg/ml gel; Amersham, Pharmacia Biotech).

Techniques: Western Blot, Clinical Proteomics, Derivative Assay, Purification, Recombinant

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Leptin and soluble leptin receptor levels in obese and weight-losing individuals.

doi: 10.1210/jcem.87.4.8381

Figure Lengend Snippet: FIG. 3. Size exclusion fractionation of plasma. Chromatographic pat- tern of plasma, following Sephacryl S300 chromatography. The two peaks correspond to sLR (f; left peak) and free leptin (E; right peak). The three capital letters correspond to the top peaks of human IgG (A; 150 kDa), human albumin (B; 60 kDa) and human L-FABP (C; 14 kDa). All fractions were assayed using human IgG, albumin, L-FABP, sLR, and leptin ELISAs.

Article Snippet: Purification of native soluble leptin receptor from plasma To purify native sLR in a single step by affinity chromatography, the murine antihuman leptin receptor mAb 2F1 was coupled to CNBrSepharose beads (2 mg/ml gel; Amersham, Pharmacia Biotech).

Techniques: Fractionation, Clinical Proteomics, Chromatography

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Leptin and soluble leptin receptor levels in obese and weight-losing individuals.

doi: 10.1210/jcem.87.4.8381

Figure Lengend Snippet: FIG. 2. ELISA for quantification of soluble leptin receptor. A, Dilu- tion-curves of plasma sample (f) and of sLR-Fc (F). Both curves show similar kinetics. B, Addition of leptin does not affect quantification of soluble leptin receptor concentration. A range of leptin concentrations was added to 100 ng/ml sLR-Fc in PBS 0.1% BSA (F) and to plasma containing 35 ng/ml native sLR (). C, Influence of leptin on the quantification of sLR was studied by making dilution curves of 7 mixtures of each two plasma samples with a high and low soluble leptin receptor level. Every point on the curves represents a mixture of the indicated ratio of dilution of both samples.

Article Snippet: Purification of native soluble leptin receptor from plasma To purify native sLR in a single step by affinity chromatography, the murine antihuman leptin receptor mAb 2F1 was coupled to CNBrSepharose beads (2 mg/ml gel; Amersham, Pharmacia Biotech).

Techniques: Enzyme-linked Immunosorbent Assay, Clinical Proteomics, Concentration Assay

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Leptin and soluble leptin receptor levels in obese and weight-losing individuals.

doi: 10.1210/jcem.87.4.8381

Figure Lengend Snippet: FIG. 4. Binding studies of 125I-leptin to sLR. Percentage binding of 125I-leptin ( 7000 cpm), added to 100 ng/ml sLR-Fc (Œ) or recombi- nant sLR-myc tagged (‚), after overnight preincubation at 4 C with different concentrations of cold leptin. Concentrations of cold leptin to sLR were in a molar ratio of respectively 0 to 1, 2:1, 6:1, and 20:1 for sLR-Fc and 0:1, 1:1, 3:1, and 10:1 for sLR-myc tagged. When 125I-leptin was added to plasma derived sLR, no binding could be observed (data not shown).

Article Snippet: Purification of native soluble leptin receptor from plasma To purify native sLR in a single step by affinity chromatography, the murine antihuman leptin receptor mAb 2F1 was coupled to CNBrSepharose beads (2 mg/ml gel; Amersham, Pharmacia Biotech).

Techniques: Binding Assay, Clinical Proteomics, Derivative Assay

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Leptin and soluble leptin receptor levels in obese and weight-losing individuals.

doi: 10.1210/jcem.87.4.8381

Figure Lengend Snippet: FIG. 5. Western blot analysis of leptin and sLR. A, Western blot analysis on the presence of leptin in plasma, on Sepharose beads coated with anti-sLR antibody 2F1 and on control Sepharose beads. After overnight incubation with plasma and carefully washing, leptin was eluted from Sepharose beads coated with anti-sLR antibodies (lane 3), indicating the presence of sLR-leptin complex bound to the beads. As expected, after incubation with plasma, no leptin was present on Sepharose beads with antirat MAC1 antibodies (lane 4), but still detectable in the plasma (lane 5). Recombinant leptin (lane 1) was used as positive control and PBS as a negative control (lane 2). In this experiment, mAb 4F8 was used for detection. B, Western blot analysis on the presence of sLR on Sepharose beads coated with leptin-PEG or coated with anti sLR antibodies. After incubation with plasma, sLR was eluted from Sepharose beads with anti sLR anti- bodies (lane 1) and from Sepharose beads with leptin-PEG (lane 2). PBS (lane 3) was used as a negative control. In this experiment, biotinylated mAb 4C3 was used for detection protection.

Article Snippet: Purification of native soluble leptin receptor from plasma To purify native sLR in a single step by affinity chromatography, the murine antihuman leptin receptor mAb 2F1 was coupled to CNBrSepharose beads (2 mg/ml gel; Amersham, Pharmacia Biotech).

Techniques: Western Blot, Clinical Proteomics, Control, Incubation, Recombinant, Positive Control, Negative Control

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Leptin and soluble leptin receptor levels in obese and weight-losing individuals.

doi: 10.1210/jcem.87.4.8381

Figure Lengend Snippet: FIG. 7. Plasma sLR and leptin concentrations in lean and morbidly obese subjects as well as in weight-losing morbidly obese individuals. Plasma concentrations of leptin (open bars) and sLR (hatched bars) in ng/ml (A) or in nanoMolar (B) in lean and morbidly obese subjects (MO). For the morbidly obese group preoperative (preop) as well as 3, 6, and 12 months postoperative (3m, 6m and 12m) values are shown. Data are depicted as mean SEM.

Article Snippet: Purification of native soluble leptin receptor from plasma To purify native sLR in a single step by affinity chromatography, the murine antihuman leptin receptor mAb 2F1 was coupled to CNBrSepharose beads (2 mg/ml gel; Amersham, Pharmacia Biotech).

Techniques: Clinical Proteomics

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Leptin and soluble leptin receptor levels in obese and weight-losing individuals.

doi: 10.1210/jcem.87.4.8381

Figure Lengend Snippet: FIG. 6. Correlation of BMI with leptin and sLR levels in a study population with a wide range of BMI. Correlation between BMI and plasma leptin levels (A) and between BMI and plasma sLR levels (B) in lean subjects and in the morbidly obese subjects before gastric restrictive surgery (n 51). The Pearson correlation coefficients are depicted in both figures.

Article Snippet: Purification of native soluble leptin receptor from plasma To purify native sLR in a single step by affinity chromatography, the murine antihuman leptin receptor mAb 2F1 was coupled to CNBrSepharose beads (2 mg/ml gel; Amersham, Pharmacia Biotech).

Techniques: Clinical Proteomics