Biomedical subjects

Charles Hanson

Publications and source records attributed to Charles Hanson.

17 recordsLinked to original sources

PubMed2006

Chromosomal integrity maintained in five human embryonic stem cell lines after prolonged in vitro culture.

There have been recent reports of human embryonic stem cell (hESC) lines developing chromosomal aberrations after long-term culture, indicating an unstable genomic status due to the in vitro milieu. This raises concern, since it would limit their use in therapeutics. In this study the chromosomal status of five well-characterized hESC lines, SA002, SA002.5, AS034.1.1, SA121 and SA461, was monitored during long-term in vitro culture. The criteria of defined hESCs were met by all of the five hESC lines (four diploid and one trisomic for chromosome 13). The genomes were screened for chromosomal aberrations and rearrangements using comparative genomic hybridization (CGH), interphase fluorescence in situ hybridization (FISH) and traditional karyotyping on several occasions while in culture. The genomic integrity was shown to be maintained after repeated freeze-thaw procedures and continuous culture in vitro for up to 22 months (148 passages). We discuss the most common de novo chromosomal aberrations reported in hESCs, as well as their possible origin.

Cell Culture Techniques↗

PubMed2006

Personality traits in women with Turner syndrome.

Turner syndrome is caused by the total or partial absence of, or structural abnormalities in, one sex chromosome. A personality style characterized by traits related to low neuroticism and high extroversion has been reported for females with Turner syndrome, indicating possible genetic factors related to personality development. This study examines personality traits in 37 Swedish women with Turner syndrome using the Karolinska Scales of Personality Inventory. Compared to reference data, the subjects rated themselves as more sensation seeking, less detached, less verbally aggressive and less irritated. These results partially support previous findings. The subjects were divided into three karyotype groups: 45,X (n = 13), 45,X/46,XX or 45,X/45,XY (n = 13), and structural abnormalities (n = 11). Contrary to some previous findings, no differences in personality were found between the karyotype groups.

Adolescent↗

PubMed2006

Donor-derived myofibroblasts in the ocular surface after allogeneic haematopoietic stem cell transplantation.

PURPOSE: To identify and characterize cells of donor origin in the ocular surface of female recipients who have undergone allogeneic haematopoietic stem cell transplantation (allo-SCT) from a male donor. METHODS: Cytological impressions from the eyes of nine allografted patients (17 eyes) were analysed. Donor cells were identified using sex-chromosome-specific fluorescence in situ hybridization (FISH). Cells were characterized by immunohistochemistry (IHC) using the CK3 and CK19 epithelial markers, the panleucocytic marker CD45 and the myofibroblast marker alpha-SMA. RESULTS: No epithelial cells of donor origin were observed in the corneal or conjunctival samples. Cells of donor origin were found in the corneal samples, although these were often too degraded to allow characterization by IHC. In the conjunctiva, a median of 86% of the total number of cells were of recipient origin, including a subgroup (2%) of giant cells exhibiting polyploidy (range 4-18 n), found in the limbal region. Donor cells were detected in the conjunctiva of all nine patients at a median ratio of 9%, of which two-thirds were CD45+/alpha-SMA+. CONCLUSIONS: We observed superficially located myofibroblasts of donor origin in all allografted patients, but not in samples from healthy controls. Whether myofibroblasts are implicated in ocular graft-versus-host disease requires further studies.

Actins↗

PubMed2005

Clonal derivation and characterization of human embryonic stem cell lines.

Human embryonic stem cells (hESC) are isolated as clusters of cells from the inner cell mass of blastocysts and thus should formally be considered as heterogeneous cell populations. Homogenous hESC cultures can be obtained through subcloning. Here, we report the clonal derivation and characterization of two new hESC lines from the parental cell line SA002 and the previously clonally derived cell line AS034.1, respectively. The hESC line SA002 was recently reported to have an abnormal karyotype (trisomy 13), but within this population of cells we observed rare individual cells with an apparent normal karyotype. At a cloning efficiency of 5%, we established 33 subclones from SA002, out of which one had a diploid karyotype and this subline was designated SA002.5. From AS034.1 we established one reclone designated AS034.1.1 at a cloning efficiency of 0.1%. These two novel sublines express cell surface markers indicative of undifferentiated hESC (SSEA-3, SSEA-4, TRA-1-60, and TRA-1-81), Oct-4, alkaline phosphatase, and they display high telomerase activity. In addition, the cells are pluripotent and form derivatives of all three embryonic germ layers in vitro as well as in vivo. These results, together with the clonal character of SA002.5 and AS034.1.1 make these homogenous cell populations very useful for hESC based applications in drug development and toxicity testing. In addition, the combination of the parental trisomic hESC line SA002 and the diploid subclone SA002.5 provides a unique experimental system to study the molecular mechanisms underlying the pathologies associated with trisomy 13.

Biomarkers↗

PubMed2005

Longterm survival of transplanted human corneal epithelial cells and corneal stem cells.

PURPOSE: To investigate the survival of donor-derived epithelial cells in conventional penetrating keratoplasty (PKP) and in homologous penetrating central limbal keratoplasty (HPCLK). METHODS AND PATIENTS: Epithelial cells from 26 eyes of 26 patients were analysed. All cases were sex-mismatched (i.e. the transplant and patient were of different genders). At suture removal more than 1 year post surgery, epithelial cells were obtained by gently wiping the removed sutures on glass slides. The cell samples were analysed using fluorescent in situ hybridization (FISH) of the sex chromosomes. This technique makes it possible to allocate the origin of each cell nucleus to either the donor or the recipient. RESULTS: All 19 conventional PKPs were clear and seven had donor-derived epithelial cells at suture removal. Five of the seven HPCLK grafts were clear at the time of investigation (365--1355 days post surgery), and donor-derived epithelial cells were found in two grafts. CONCLUSION: Harvesting cells from removed sutures in combination with FISH enables the clinical study of cell survival in corneal transplants without jeopardizing functioning grafts. From the limited sample investigated, the following tentative conclusions can be made. Donor-derived epithelial cells can remain in conventional PKP for over 1 year. In combined stem cell and corneal grafts (HPCLK), donor-derived epithelial cells may also be retrieved at 1 year or beyond following surgery but the correlation between their presence and a remaining clear graft is uncertain.

Adult↗

PubMed2005

Longterm survival of transplanted human corneal epithelial cells and corneal stem cells.

PURPOSE: To investigate the survival of donor-derived epithelial cells in conventional penetrating keratoplasty (PKP) and in homologous penetrating central limbal keratoplasty (HPCLK). METHODS AND PATIENTS: Epithelial cells from 26 eyes of 26 patients were analysed. All cases were sex-mismatched (i.e. the transplant and patient were of different genders). At suture removal more than 1 year post surgery, epithelial cells were obtained by gently wiping the removed sutures on glass slides. The cell samples were analysed using fluorescent in situ hybridization (FISH) of the sex chromosomes. This technique makes it possible to allocate the origin of each cell nucleus to either the donor or the recipient. RESULTS: All 19 conventional PKPs were clear and seven had donor-derived epithelial cells at suture removal. Five of the seven HPCLK grafts were clear at the time of investigation (365-1355 days post surgery), and donor-derived epithelial cells were found in two grafts. CONCLUSION: Harvesting cells from removed sutures in combination with FISH enables the clinical study of cell survival in corneal transplants without jeopardizing functioning grafts. From the limited sample investigated, the following tentative conclusions can be made. Donor-derived epithelial cells can remain in conventional PKP for over 1 year. In combined stem cell and corneal grafts (HPCLK), donor-derived epithelial cells may also be retrieved at 1 year or beyond following surgery but the correlation between their presence and a remaining clear graft is uncertain.

Adult↗

PubMed2004

Hypothyroidism is common in turner syndrome: results of a five-year follow-up.

Turner syndrome (TS) is caused by a sex chromosome aberration. The aim was to study the prevalence and incidence of thyroid disease in adults with TS. Women with TS (n = 91; mean age, 37.7 +/- 11 yr) were compared with an age-matched female random population sample (n = 228). At baseline, 15 (16%) TS women were treated for hypothyroidism, and elevated serum TSH was found in another eight (9%). As a result, hypothyroidism was more common in women with TS (25%) than in controls (2%; P < 0.0001). Serum free T4 was lower (P = 0.02), and serum TSH was higher (P < 0.0001) in TS women than in age-matched controls. Of all TS women with hypothyroidism, 10 (43%) had an elevated thyroid peroxidase antibody titer vs. 15 (22%) of those without hypothyroidism (P < 0.05), evenly distributed between the karyotype 45,X and mosaicism. A high body mass index, but not a family history or blood lipids, was associated with hypothyroidism in TS. After the 5-yr follow-up, an additional 11 (16%) developed hypothyroidism, of whom four (36%) had elevated thyroid peroxidase. Altogether, 34 (37%) TS women had hypothyroidism after the 5-yr follow-up. Autoimmune hypothyroidism was common, with an annual incidence of 3.2% in TS. Thyroid function should be checked regularly in TS.

Adolescent↗

PubMed2004

Spontaneous pregnancies in a Turner syndrome woman with Y-chromosome mosaicism.

PURPOSE: To present a case involving pregnancies in a Turner woman with Y-chromosome mosaicism. METHOD: A descriptive case report of a single patient. RESULTS: A 39-year-old woman was admitted to the endocrine clinic due to fatigue and premature menopause. She had tried in-vitro fertilization and oocyte donation twice without pregnancies but became spontaneously pregnant at age 36 and 37 and delivered two girls. During the seventh month of the second pregnancy, a dissecting aortic aneurysm, a coarctation, and subsequently a pheochromocytoma were detected and repaired. Hypothyroidism developed. Turner syndrome was diagnosed. Fluorescence in situ hybridization (FISH) analysis of lymphocytes revealed 31% XY cells and 4% XYY cells, while 66% of buccal cells had an XY constitution. Oophorectomy revealed no malignancy. FISH revealed 54% XY cells in the left gonad and 38% XY cells in the right. CONCLUSION: Turner syndrome should be suspected in women with aortic dissection, in general, but especially in those with additional features such as horseshoe kidney, coarctation, and infertility.

Adult↗

PubMed2004

Time-lapse recordings of human corneal epithelial healing.

PURPOSE: The aim of this study was to design an experimental set-up for the study of human corneal epithelial wound healing in a controlled in vitro situation. METHODS: A time-lapse set-up was used. This allowed for pictures to be captured with a magnification ranging from x 80 to x 1800. Pictures were captured at 1-min intervals during the observation period, which lasted up to 4 days. Human corneal tissue was obtained from the Eye Bank or from surgery. A small, rounded lesion was produced in the corneal epithelium with a miniature drill. The specimens were placed in a mini-incubator; the camera focused on the epithelial lesion and continuously observed using the time-lapse set-up. RESULTS: The healing process of human corneal epithelium could be followed for several days. The initial healing response could be divided into a slow, a rapid and a consolidating phase. The first two phases lasted about 12 hours, and by then, epithelial cells covered the lesion. Depending on the origin of the tissue and the placement of the lesion, variations in the healing response could be seen. CONCLUSION: The time-lapse technique makes it possible to study epithelial wound healing over time at the cellular level. Data collected in this way can fill the gap between in vivo studies, where, by nature, human wound healing studies are restricted, and cell culture techniques, where cellular responses in many cases differ from the in vivo situation.

Epithelium, Corneal↗

PubMed2004

Derivation, characterization, and differentiation of human embryonic stem cells.

The derivation of human embryonic stem (hES) cells establishes a new avenue to approach many issues in human biology and medicine for the first time. To meet the increased demand for characterized hES cell lines, we present the derivation and characterization of six hES cell lines. In addition to the previously described immunosurgery procedure, we were able to propagate the inner cell mass and establish hES cell lines from pronase-treated and hatched blastocysts. The cell lines were extensively characterized by expression analysis of markers characteristic for undifferentiated and differentiated hES cells, karyotyping, telomerase activity measurement, and pluripotency assays in vitro and in vivo. Whereas three of the cell lines expressed all the characteristics of undifferentiated pluripotent hES cells, one cell line carried a chromosome 13 trisomy while maintaining an undifferentiated pluripotent state, and two cell lines, one of which carried a triploid karyotype, exhibited limited pluripotency in vivo. Furthermore, we clonally derived one cell line, which could be propagated in an undifferentiated pluripotent state.

Animals↗

PubMed2003

[Cloning techniques for mitochondrial diseases and prenatal diagnosis].

Disorders caused by mutation in the mitochondrial DNA are uncommon. Due to the special pattern of inheritance and of the variability of penetrance the options to affected couples to have healthy children are few. So far traditional prenatal diagnosis is of limited benefit. The problems may be overcome by oocyte donation. However, if the couple wants their own biological offspring, no good method is available today. We discuss ooplasmic and nuclear transfer as possible future options for these couples to have healthy biological children.

Child↗

PubMed2003

A morphological and chromosomal study of blastocysts developing from morphologically suboptimal human pre-embryos compared with control blastocysts.

BACKGROUND: IVF laboratories performing embryo transfer at day 2 or 3 after fertilization are currently discarding pre-embryos considered suboptimal using morphological criteria. The objective of this study was to investigate whether blastocysts, cultured from such pre-embryos (surplus), were chromosomally and morphologically normal. As a control group we used morphologically good quality embryos (GQE), cultured to the blastocyst stage. METHODS: Human pre-embryos considered suboptimal were cultured to the blastocyst stage. As a control group, frozen-thawed pre-embryos of good quality were cultured under identical conditions. The chromosomal status of the blastocysts obtained was studied by multi-colour fluorescence in-situ hybridization for chromosomes 13, 16, 18, 21, 22, X and Y. RESULTS: There is, on average, a significantly higher degree of chromosomal aberrations in blastocysts derived from surplus pre-embryos compared to blastocysts derived from GQE, and the chromosomal aberrations are generally found in a higher number of blastomeres per blastocyst. In addition, blastocysts from surplus pre-embryos had significantly poorer morphology compared to GQE. Improvement in morphology and/or developmental rate in surplus pre-embryos between day 2 and day 3 did not predict a morphologically/chromosomally normal blastocyst. However, this study shows that close to half of the surplus pre-embryos that reach the blastocyst stage can be considered chromosomally normal when assessed for these seven chromosomes. Furthermore, we found that chromosomal aberrations were more concentrated in a particular cell population within blastocysts derived from GQE, compared with surplus blastocysts. CONCLUSIONS: The study suggests that even if the IVF laboratory is on average making the correct decision about the potential of a pre-embryo, surplus pre-embryos that might become chromosomally normal blastocysts are still being discarded.

Adult↗

PubMed2002

Is any form of gender selection ethical?

In later years, sex selection has become of importance for prevention of X-linked diseases in families at risk. There is today a potential to perform sperm selection before fertilization by taking advantage of the chromosomal heterogamy of spermatozoa, and before implantation by preimplantation genetic diagnosis (PGD). The methods of sex determination by separating sperrmatozoa are, in our opinion, still not safe enough for routine clinical use. Apart from the technical problems and possible associated risks, which first must be better evaluated, the most critical questions are ethical or legal. We support the use of sex selection by PGD in X-linked severe disease, but due to the potential risks of misuse, we are not prepared to support a more liberal attitude as long as the discriminated sex in nearly all parts of the world are women.

Female↗

PubMed2002

Survival of transplanted human corneal stem cells. Case report.

PURPOSE: To report a case history involving long-term survival of transplanted human corneal stem cells. METHODS: A male patient with severe bilateral chemical burns received six corneal transplants, all of which failed. He subsequently received combined corneal transplants and stem cell transplants, which have remained clear for 3 and 4 years respectively. One of the donors was female. We studied the gender of the epithelial cells of the cheek of the patient and of the two grafts using fixation and fluorescent in situ hybridization (FISH) analyses. RESULTS: In the graft from the female donor, 30% of the epithelial cells were of female origin. All the epithelial cells from the cheek and the other graft were of male origin. CONCLUSION: This case demonstrates that transplanted human corneal stem cells can survive and replicate in the long-term (3 years) without systemic immunosuppression. The case also indicates that a minority (30%) of healthy transplanted epithelial cells is enough to present a clear graft with a clinically healthy ocular surface.

Aged↗

PubMed2002

Fluorescence in situ hybridisation analysis and ovarian histology of women with Turner syndrome presenting with Y-chromosomal material: a correlation between oral epithelial cells, lymphocytes and ovarian tissue.

The early detection of Y-chromosomal material in women with Turner syndrome (TS) is of great importance due to a relatively high risk of gonadal tumour development. Using fluorescence in situ hybridisation (FISH) analysis, we studied the presence of three different Y-specific sequences (SRY, Ycen and Yq12) in three different tissues (oral epithelial cells, lymphocytes and ovarian tissue) of twelve TS women. We have also described their ovarian histology. Two of the women (17%) had gonadal tumours. In five women where ovarian tissue was available, the presence of Y-chromosomal material in oral epithelial cells and lymphocytes correlated to the presence of Y-chromosomal material in the gonads. We therefore conclude that FISH analysis of oral epithelial cells and/or lymphocytes is a valuable complement to karyotyping for the early detection of Y-chromosomal material in TS women.

Adult↗

PubMed2002

A human oocyte with two sets of MII/PB-structures.

This paper reports a case where a normal-sized human oocyte has been documented having two polar bodies and two metaphase spindles. This finding suggests that the chromosomal content in oocytes of normal size may occasionally be duplicated and offers an additional explanation for the origin of triploid zygotes in humans. We speculate that this duplication may be caused by oocyte fusion at an early stage.

Adult↗

PubMedDate not supplied

Human embryonic stem cells and chromosome stability.

The use of human embryonic stem cells (HESC) in research is increasing exponentially and HESC will certainly be of importance in biological, clinical and toxicological research for many years to come. Once established, HESC lines are expected to be chromosomally stable. However, our own experience of culturing HESC and some published reports indicate that HESC may show chromosomal instability while being cultured continuously in vitro. We conclude that the effects of different culture techniques and long-term culture on the chromosome stability of HESC still remain to be elucidated and we recommend regular analysis of the chromosome constitution in cell lines using traditional karyotyping, CGH, FISH and PCR. We also recommend freezing of HESC at low passage number and in larger batches after thawing and expansion in order to secure material in case mutations occur in the cell line at a later stage of culture.

Cell Culture Techniques↗