OBJECTIVE: We investigated whether the insemination method (in vitro fertilization [IVF] or intracytoplasmic sperm injection [ICSI]) affected morphokinetic events and abnormal cleavage events in embryonic development. METHODS: A total of 1,830 normal fertilized embryos were obtained from 272 IVF and ICSI cycles that underwent ovum retrieval culture using a time-lapse system (Embryoscope) from June 2013 to March 2015. All embryos were investigated by a detailed time-lapse analysis that measured the developmental events in the hours after IVF or ICSI insemination. RESULTS: No significant differences were observed between the two groups regarding clinical outcomes (p>0.05). ICSI-derived embryos showed significantly faster morphokinetics than those derived from conventional IVF, from the time to pronuclear fading to the time to 6 cells (p<0.05). However, no significant differences were found from the time to 7 cells to the time to expanded blastocyst (p>0.05). There were no differences in abnormal cleavage events between the two groups (p>0.05); they showed the same rates of direct cleavage from 1 to 3 cells, 2 multinucleated cells, 2 uneven cells, and reverse cleavage. CONCLUSION: The morphokinetics of embryo development was found to vary between IVF- and ICSI-fertilized oocytes, at least until the 6-cell stage. However, these differences did not affect the clinical outcomes of the embryo. Additionally, no significant differences in abnormal cleavage events were found according to the fertilization method.
Blastocyst ; Embryonic Development* ; Embryonic Structures* ; Female ; Fertilization ; Fertilization in Vitro ; Humans* ; In Vitro Techniques* ; Insemination ; Methods ; Oocytes* ; Ovum ; Pregnancy ; Sperm Injections, Intracytoplasmic ; Spermatozoa* ; Time-Lapse Imaging

OBJECTIVE: Indocyanine green with near-infrared fluorescence imaging (NIR-ICG) is a new tracer modality in the limelight used for lymphatic mapping. The advantage of this method is to provide real-time image during surgery. To use ICG for image guided lymph node dissection, a surgeon needs to know initial appearing time and duration. METHODS: A 52-year-old woman undertook surgery diagnosed with endometrial cancer. She had no past medical history and her body mass index was 25.3 kg/m2. Preoperative magnetic resonance imaging examination revealed 2.7 cm sized cancerous mass in the endometrial cavity with superficial myometrial invasion without lymph node enlargement. Four mL (1.25 mg/mL) of ICG solution was prepared for injection. For each site, 1 mL of solution was injected superficially, 2-3 mm into the cervical submucosa and another 1 mL was injected deep, 1-2 cm into the stroma of the cervix. We recorded video with 30° 10 mm scope equipped with a specific lens and light source emitting both visible and NIR light (KARL STORZ GmbH & Co. KG, Tuttlingen, Germany). RESULTS: Pelvic lymph node was visualized from around 5 minutes. ICG was dispersed into organs after hysterectomy (53 minutes after ICG injection), yet we could clearly identify sentinel lymph node (SLN). Pathology revealed endometriod adenocarcinoma grade I, myometrial invasion with less than half of myometrium and no lymph node metastasis. CONCLUSION: Cervical injection of ICG provides good visualization of SLN from 5 minutes to over an hour. Our film gives an idea about time management to make a plan for surgery and not to miss SNLs.
Adenocarcinoma/diagnostic imaging/*pathology ; Coloring Agents/*therapeutic use ; Endometrial Neoplasms/diagnostic imaging/*pathology ; Female ; Humans ; Indocyanine Green/*therapeutic use ; Lymphatic Metastasis/diagnostic imaging ; Middle Aged ; Optical Imaging/methods ; Sentinel Lymph Node/*diagnostic imaging ; Time-Lapse Imaging/methods

Molecular mechanisms underlying the effects of Fyn on cell morphology, pseudopodium movement, and cell migration were investigated. The Fyn gene was subcloned into pEGFP-N1 to produce pEGFP-N1-Fyn. Chinese hamster ovary (CHO) cells were transfected with pEGFP-N1-Fyn. The expression of Fyn mRNA and proteins was monitored by reverse transcription-PCR and Western blotting. Additionally, transfected cells were stained with 4',6-diamidino-2-phenylindole and a series of time-lapse images was taken. Sequences of the recombinant plasmids pMD18-T-Fyn and pEGFP-N1-Fyn were confirmed by sequence identification using National Center for Biotechnology Information in USA, and Fyn expression was detected by RT-PCR and Western blotting. The morphology of CHO cells transfected with the recombinant vector was significantly altered. Fyn expression induced filopodia and lamellipodia formation. Based on these results, we concluded that overexpression of mouse Fyn induces the formation of filopodia and lamellipodia in CHO cells, and promotes cell movement.
Animals ; Blotting, Western ; CHO Cells ; Cricetinae ; Cricetulus ; Genetic Vectors ; Green Fluorescent Proteins/genetics ; Mice ; Proto-Oncogene Proteins c-fyn/genetics/*metabolism ; Pseudopodia/*metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Time-Lapse Imaging ; Transfection

B lymphocyte cell senses and acquires foreign antigens through clonal distributed B cell receptors (BCRs) expressed on the surface of plasma membrane. The presentation formats of antigens are quite diverse. Based on their Brownian diffusion mobility, there are three forms: free mobile soluble antigens, lateral mobile membrane bound antigens, and fixed immobile antigens. Here, using high resolution high speed live cell imaging approaches, we provide evidence that BCR microclusters are formed on the surface of B cells shortly after B cell's encountering of antigens with each format of motion features. Through high speed live cell imaging, we determine that these BCR microclusters show dynamic growth feature and by doing so function as the basic platforms for B cells to acquire the antigens. We propose that the formation and dynamic growth of BCR microcluster is a universal mechanism for B cell to response to antigens with diverse motion features.
Adaptive Immunity ; Animals ; Antigens ; immunology ; metabolism ; B-Lymphocytes ; immunology ; metabolism ; Cell Membrane ; metabolism ; Cells, Cultured ; Humans ; Mice ; Mice, Transgenic ; Microscopy, Fluorescence ; Protein Transport ; immunology ; Receptors, Antigen, B-Cell ; metabolism ; Single-Cell Analysis ; Time-Lapse Imaging