Biomedical subjects
Publications and source records attributed to M Ginsburg.
HLA-G is believed to play a pivotal role in the immunoprotection of the semiallogenic embryo. Its expression during pre- and early implantation is correlated with the cleavage rate of the embryo. Studies in congenic mice have revealed that mRNA of both the maternal and paternal haplotypes are present in zygotes and in embryos at all stages of development.
BACKGROUND: We have previously reported the retrospective observation that when at least one embryo, transferred on day 3, expressed sHLA-G above the geometric mean (sHLA-G+) 46 h post-ICSI, there was a marked improvement in both pregnancy (PR) and implantation (IR) rates. METHODS: The media surrounding individual embryos derived from ICSI performed on oocytes from 482 women < or =43 years of age were tested for sHLA-G expression by specific ELISA. RESULTS: We report here prospective results showing improved IVF results following the transfer of 'good quality' embryos (7-9 cells with <20% fragmentation) by preferentially including at least one sHLA-G+ embryos. PR and IR for women < or =38 years were 63% and 32% when one transferred embryo was sHLA-G+, and 69% and 36% when at least two embryos were sHLA-G+. When none of the embryos transferred was sHLA-G+, PR and IR were 25% and 13%, respectively. Comparable PR and IR for women 39-43 years were 29% and 11% when none of the transferred embryos were sHLA-G+; 38% and 15% when at least one sHLA-G+ embryo was transferred; and 61% and 26% when at least two 2 sHLA-G+ embryos were transferred. The data were stratified by patient age. CONCLUSIONS: PR and IR increased with the addition of each sHLA-G+ embryo, regardless of age. While there are significant barriers to routine embryo sHLA-G testing, we believe that if implemented, this would provide a mechanism for optimizing IVF PR while minimizing the risk of multiple pregnancies.
Non-mammalian vertebrate erythrocytes are flattened nucleated ellipsoids containing marginal bands (MBs) of microtubules that assemble during cellular morphogenesis. Earlier work suggested that pointed erythroid cells containing pointed MBs were intermediate stages in terminal differentiation, rather than aberrant forms, but direct evidence was lacking. Here we report on morphogenesis in individual post-cytokinetic amphibian erythroblasts in culture. Daughter cells remained adjacent in pairs, and developed pointed morphology over 1-2 h in the following sequence: (a) ends opposite the cytokinetic furrow became pointed, producing a spheroidal singly-pointed stage; (b) furrow ends usually became pointed, yielding doubly-pointed cells; (c) furrow-end points disappeared, producing a second singly-pointed stage that was flattening. Over a longer term, the single points sometimes disappeared, yielding a flattened discoid. These observations support the hypothesis that pointed cells are normal intermediates in a biogenetic program in which post-mitotic centrosomes organize MBs while occupying the singly-pointed ends of differentiating erythroblasts.
Pulmonary emphysema is believed to result from an imbalance between proteolytic enzymes and their inhibitors. Multiple studies have examined the presence of various proteases within the bronchoalveolar lavage fluid from patients with chronic obstructive pulmonary disease (COPD). However, to date extensive examination of the lung parenchyma for the expression of destructive enzymes has not yet been determined. The following study examines the lung parenchyma of 23 patients with emphysema and 8 normal control samples for the expression of matrix matalloproteinase-1 (MMP-1), MMP-12, and MMP-9. We report here that interstitial collagenase (MMP-1) RNA, protein, and activity are present in the lung parenchyma of patients with emphysema and not in the lung of normal control subjects. In contrast, metalloelastase (MMP-12) expression is absent in these samples. Immunohistochemistry studies localized MMP-1 to the Type II pneumocyte in patients with emphysema and not normal control subjects or smokers without emphysema. This observation demonstrates that the lung is altered in emphysema such that the Type II pneumocyte secretes MMP-1 and suggests that MMP-1 may be an important enzyme involved in the destruction of the lung in the human disease. In addition, the induction of a proteolytic enzyme within the Type II pneumocyte suggests that the cells within the lung itself are capable of producing degradative enzymes in this disease process.
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The purpose of this investigation was to determine whether there were differences in the magnitude of insular cortex activation across varying intensities of static and dynamic exercise. Eighteen healthy volunteers were studied: eight during two intensities of leg cycling and ten at different time periods during sustained static handgrip at 25% maximal voluntary contraction or postexercise cuff occlusion. Heart rate, blood pressure (BP), perceived exertion, and regional cerebral blood flow (rCBF) distribution data were collected. There were significantly greater increases in insular rCBF during lower (6.3 +/- 1.7%; P < 0.05) and higher (13.3 +/- 3.8%; P < 0.05) intensity cycling and across time during static handgrip (change from rest for right insula at 2-3 min, 3.8 +/- 1.1%, P < 0.05; and at 4-5 min, 8.6 +/- 2.8%, P < 0.05). Insular rCBF was decreased during postexercise cuff occlusion (-5.5 +/- 1.2%; P < 0.05) with BP sustained at exercise levels. Right insular rCBF data, but not left, were significantly related, with individual BP changes (r(2) = 0.80; P < 0.001) and with ratings of perceived exertion (r(2) = 0.79; P < 0.01) during exercise. These results suggest that the magnitude of insular activation varies with the intensity of exercise, which may be further related to the level of perceived effort or central command.
The origin of the germline is studied in avians by tracing primordial germ cells from the stage of the germinal crescent backwards to earlier developmental stages. It has been demonstrated that during primitive streak formation, the germline has already been segregated. However, during this stage, the cells are seen gradually migrating from the epiblast to the hypoblast. The vertical migration is followed by a horizontal translocation to the extra-embryonic germinal crescent, being carried out by the hypoblast that is pushed anteriorly by the invading endodermal cells. In contrast, it has been shown that in the mouse the germ cells are allocated during gastrulation in a cluster of cells. Our results demonstrate that in avians the allocation takes place according to a different mode. The close association between the germ cells and the extra-embryonic mesoderm indicates that the germline in avians, as in the mouse, develops from a subset of cells that have already segregated from the epiblast as extra-embryonic mesoderm.
The origin of the germ cell lineage in vertebrates is a fundamental question that has preoccupied developmental biologists. Recent work on the origin of the avian germ line has extended and clarified our understanding of the temporal and spatial segregation of primordial germ cells (PGC) during prestreak stages of development. The germ cells first appear at Stage X (Eyal-Giladi and Kochav, 1976) in the ventral surface of the area pellucida in a scattered pattern among polyingressing cells. Subsequently, the PGC gradually translocate from the epiblast to the hypoblast. The entire process appears to be dependent upon the maintenance of an organized area pellucida. Little is known about the regulatory events governing germ cell emergence during this period; however, the culture of dispersed blastodermal cells on a mouse fibroblast feeder layer can compensate for a disorganized area pellucida and offers an in vitro system to examine the molecular basis of germ cell development. Such basic information is valuable for current approaches towards the production of transgenic poultry with targeted changes to the genome through the use of avian embryonic stem cells or primordial germ cells. Refinement of the culture of primordial germ cells or their precursors should allow academic and industrial research laboratories to answer significant biological questions and to improve the genetic potential of commercial poultry stocks. A better understanding of the biology of avian primordial germ cells during early embryo development can only enhance this process.
The temporal and spatial pattern of segregation of the avian germline from the formation of the area pellucida to the beginning of primitive streak formation (stages VII-XIV, EG&K) was investigated using the culture of whole embryos and central and peripheral embryo fragments on vitelline membranes at stages VII-IX, immunohistological analysis of whole mount embryos and sections with monoclonal antibodies MC-480 against stage-specific embryonic antigen-1 (SSEA-1) and EMA-1, and with the culture of dispersed blastoderms at stages IX-XIV with and without on STO feeder layer. Whole embryos at intrauterine stages developed up to the formation of the primitive streak despite the absence of area pellucida expansion. Primordial germ cells (PGCs) appeared in the cultures of whole embryos and only in central fragments containing a partially formed area pellucida at stages VII-IX. When individual stage IX-XIV embryos were dispersed and cultured without a feeder layer, 25-45 PGCs/embryo were detected only with stage X-XIV, but not with stage IX blastoderms. However, the culture of dispersed cells from the area pellucida of stages IX-XIII on STO feeder layers yielded about 150 PGCs/embryo. The carbohydrate epitopes recognized by anti-SSEA-1 and EMA-1 first appeared at stage X on cells in association with polyingressing cells on the ventral surface of the epiblast and later on the dorsal surface of the hypoblast. The SSEA-1-positive hypoblast cells gave rise to chicken PGCs when cultured on a feeder layer of quail blastodermal cells. From these observations, we propose that the segregation and development of avian germline is a gradual, epigenetic process associated with the translocation of SSEA-1/EMA-1-positive cells from the ventral surface of the area pellucida at stage X to the dorsal side of the hypoblast at stages XI-XIV.
BACKGROUND: Cerebral injury remains a significant complication of cardiac operations. We determined the incidence of cerebral dysfunction in a population of elderly patients undergoing open chamber cardiac operations (group 1) as compared with a younger population (group 2) and an age-matched group of elderly patients undergoing major noncardiac operations (group 3). METHODS: Sixty-eight patients (55 for open chamber cardiac operations and 13 for noncardiac operations) were prospectively studied. Patients were evaluated preoperatively and postoperatively before hospital discharge using a complete neurologic examination and a battery of standard neuropsychometric tests, and at surgical follow-up with neuropsychometric tests only. RESULTS: Postoperative changes detected by neurologic examination consisted of the appearance of new primitive reflexes in all groups. No statistically significant differences in incidence were found. The neuropsychometric performance of group 1 patients was statistically different from that of patients in groups 2 and 3 only in the early follow-up period. CONCLUSIONS: Elderly patients having open chamber cardiac operations exhibit significantly more cerebral dysfunction in the early postoperative period than those undergoing major noncardiac operations and younger patients after open chamber procedures. These changes do not persist into the late follow-up period.
At the early somite stage, chicken embryo primordial germ cells comprise a population of about 200 dispersed, morphologically distinct cells, located in the extraembryonic germinal crescent. Tracing the origin of primordial germ cells at earlier stages is at present not possible by either cytological or immunohistochemical techniques. To resolve this difficulty we have microdissected embryos at early embryonic stages, isolated fragments from various positions and incubated them to the somite stage, when the primordial germ cells can be visualized accurately. From these studies a distinct developmental pattern has emerged. (1) At the uterine stage, when the area pellucida starts to thin out, there is already a defined area, confined to the most central part of the blastodisc, which constitutes the sole source of the future primordial germ cells. (2) As development proceeds, the area that gives rise to the primordial germ cells expands peripherally. (3) Primordial germ cells thereafter migrate as isolated cells, descending to the lower layer, the hypoblast. (4) This gradual process starts at the onset of blastulation and terminates towards the end of gastrulation. (5) The hypoblast, destined to become the yolk sac endoderm, is pushed anteriorly during gastrulation by morphogenetic movements to be replaced by the endoderm proper; the primordial germ cells are carried forwards with it and occupy an extraembryonic location.
OBJECTIVES: This study attempted to document the incidence of pulmonary vein complications and their potential relation to clinical outcome in patients after lung transplantation. BACKGROUND: Several case reports have documented the presence of pulmonary venous thrombosis causing graft failure in patients after lung transplantation. Because the presentation of these complications mimics that of other postoperative problems, the true incidence of pulmonary vein abnormalities remains unclear. Transesophageal echocardiography is ideally suited to examine the pulmonary veins in the postoperative setting. METHODS: Twenty-one consecutive patients undergoing lung transplantation at our institution underwent transesophageal echocardiography within 32 days of transplantation (mean [+/- SD] 6.5 +/- 7.8 days). Special attention was placed on visualizing the pulmonary veins. RESULTS: Six (29%) of the 21 patients were noted to have abnormalities of the pulmonary veins in the vicinity of the anastomotic site. After follow-up of 30 days, 4 of these patients (67%) had significant cardiovascular morbidity, and 2 died, compared with 1 (7%) of 15 patients with normal pulmonary veins (p = 0.03). The degree of obstruction of the pulmonary vein appeared to correlate with short-term outcome. CONCLUSIONS: Abnormalities of the pulmonary veins are common after lung transplantation and are easily identified by transesophageal echocardiography. Occlusive thrombi appear to be detrimental to short-term outcome.
Sax1 (previously CHox3) is a chicken homeobox gene belonging to the same homeobox gene family as the Drosophila NK1 and the honeybee HHO genes. Sax1 transcripts are present from stage 2 H&H until at least 5 days of embryonic development. However, specific localization of Sax1 transcripts could not be detected by in situ hybridization prior to stage 8-, when Sax1 transcripts are specifically localized in the neural plate, posterior to the hindbrain. From stages 8- to 15 H&H, Sax1 continues to be expressed only in the spinal part of the neural plate. The anterior border of Sax1 expression was found to be always in the transverse plane separating the youngest somite from the yet unsegmented mesodermal plate and to regress with similar dynamics to that of the segregation of the somites from the mesodermal plate. The posterior border of Sax1 expression coincides with the posterior end of the neural plate. In order to study a possible regulation of Sax1 expression by its neighboring tissues, several embryonic manipulation experiments were performed. These manipulations included: removal of somites, mesodermal plate or notochord and transplantation of a young ectopic notochord in the vicinity of the neural plate or transplantation of neural plate sections into the extraembryonic area. The results of these experiments revealed that the induction of the neural plate by the mesoderm has already occurred in full primitive streak embryos, after which Sax1 is autonomously regulated within the spinal part of the neural plate.
Most surgeons consider patients with solitary adrenal metastasis from a primary lung carcinoma incurable and avoid excision of both the adrenal and primary lung tumors. However, several cases of successful surgical management of these patients recently have been reported. We reviewed 12 surgically treated patients with isolated adrenal and lung disease and identified 2 survivors of greater than fifteen years (17%) and 4 additional patients who are still alive following combined resection (34%). This survival rate, albeit in a selected population, represents an improvement over the natural history of nine months' survival. We suggest that if after six to twelve months of following patients with lung cancer and isolated adrenal metastasis no other evidence of spread of disease is evident, the tumor biology may be favorable and resection of both adrenal and lung lesions is reasonable.
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The enzyme-AMP reaction intermediate of the 102-kDa bovine DNA ligase I was digested with trypsin, and the adenylylated peptide was isolated by chromatography under conditions that maintain the acid-labile phosphoramidate bond. Microsequencing of the peptide showed that it contains an internal trypsin-resistant lysine residue, as expected for the site of adenylylation. Inhibition of DNA ligase I activity by pyridoxal 5'-phosphate also indicated the presence of a reactive lysine residue in the catalytic domain of the enzyme. Comparison of the known primary structures of several other DNA ligases with the adenylylated region of mammalian DNA ligase I allows their active sites to be tentatively assigned by sequence homology. The ATP-dependent DNA ligases of mammalian cells, fission yeast, budding yeast, vaccinia virus, and bacteriophages T3, T4, and T7 contain the active site motif Lys-Tyr/Ala-Asp-Gly-(Xaa)-Arg, with the reactive lysine residue flanked by hydrophobic amino acids. The distance between the postulated adenylylation site and the carboxyl terminus of the polypeptide is very similar in these ATP-dependent DNA ligases, whereas the size of the amino-terminal region is highly variable.
The open reading frame corresponding to BORF2 and encoding the large subunit of the Epstein-Barr virus (EBV) ribonucleotide reductase has been inserted into the prokaryotic expression vector pUC19. A 90-kDa protein was produced when the intact plasmid was used as a template for in vitro DNA-directed protein synthesis. Using templates generated by restriction digests within the BORF2 open reading frame, in the same system, truncated polypeptides resulted confirming the identity of the 90-kDa protein. The protein was then produced in a heterologous expression system and used in protein immunoblotting to screen for antibodies in sera from nasopharyngeal carcinoma (NPC), Burkitt's lymphoma (BL) or control subjects. Twenty out of 33 NPC sera were positive for antibodies against the large subunit, 13 of these were positive for both IgG and IgA, whilst 7 were positive for IgG only. Out of 15 BL sera and 10 control sera, none were positive. These results are similar to those observed for other EBV-encoded enzymes, including the DNase which had been used as an early marker for the development of NPC. The results presented here indicate that antibodies against the large subunit of ribonucleotide reductase could serve as an additional marker for NPC.
With the aid of a whole-mount technique, we have detected a small cluster of alkaline phosphatase (ALP)-positive cells in whole mounts of mid-primitive-streak-stage embryos, 7-7 1/4 days post coitum (dpc). Within the cluster, about 8 cells contain a small cytoplasmic spot, intensely stained for ALP activity and possibly associated with an active Golgi complex. The cluster lies just posterior to the definitive primitive streak in the extraembryonic mesoderm, separated from the embryo by the amniotic fold. Towards the end of gastrulation, the number of cells containing the ALP-positive spot rises to between 50 and 80. Thereafter the number of cells in the extraembryonic cluster declines, and similar cells start to be seen in the mesoderm of the primitive streak and then in the endoderm. At 8 dpc, about 125 ALP-stained cells are found, mainly in the hindgut endoderm and also at the base of the allantois, their appearance and location at this stage agreeing closely with previous reports on primordial germ cells (PGCs). Embryos from which the cluster area has been removed at the 7-day stage are devoid of PGCs after culture for 48 h, whereas the excised tissue is rich in PGCs. We argue that the cells in the cluster are indeed primordial germ cells, at a stage significantly earlier than any reported previously. This would indicate that the PGC lineage in the mouse is set aside at least as early as 7 dpc, possibly as one of the first 'mesodermal' cell types to emerge, and that its differentiation, as expressed by ALP activity, is gradual.