Successful ovarian tissue cryopreservation with transvaginal natural orifice transluminal endoscopic surgery: A case report.

Chemotherapy and radiation therapy can cause gonadal dysfunction in women of reproductive age. Ovarian tissue cryopreservation is performed to restore fertility by allowing transplantation of the patient's frozen-thawed ovarian tissue or through future in vitro maturation and in vitro fertilization of frozen-thawed oocytes. Herein, we describe our initial experience with vaginal natural orifice transluminal endoscopic surgery for ovarian tissue preservation in a young woman with malignant tumor. A 23-year-old woman with anaplastic lymphoma kinase-positive malignant lymphoma was scheduled for hematopoietic stem cell transplantation after experiencing relapse following R-cyclophosphamide, doxorubicin, vincristine, and prednisolone therapy. Ovarian tissue cryopreservation was selected as only MII2 oocytes were collected. Vaginal natural orifice transluminal endoscopic surgery was performed to excise the left ovary. Ovarian tissues were frozen using the vitrification method. The operative time was 37 min, and blood loss was minimal. Pathological examination revealed no metastatic findings of malignant lymphoma and no thermal damage to the ovarian tissue due to bipolar disorder. The patient was discharged on the first day postoperatively, and her postoperative course was uneventful. The vaginal natural orifice transluminal endoscopic surgery technique can provide a safe and effective alternative to laparoscopy or laparotomy for the cryopreservation of ovarian tissue in young patients with cancer. We believe this method has potential application in sexually mature female cancer survivors.

Ovarian tissue cryopreservation with vaginal natural orifice transluminal endoscopic surgeryChemotherapy and radiotherapy can affect a woman's ability to have children by reducing ovarian function. This can make it hard to conceive even with fertility treatments. Freezing healthy ovaries before these treatments can help restore fertility. This can be done by freezing and later transplanting ovarian tissue or by fertilizing frozen eggs in a lab. Traditional surgery to remove ovaries can cause cosmetic issues and pain. But now, a new method called vaginal spontaneous opening transperitoneal endoscopic surgery is becoming more common. This surgery is less invasive, quicker, and causes less bleeding. We recently used this method to preserve ovarian tissue in young women with cancer. The surgery was successful with minimal complications. This new approach could offer a safer option for preserving fertility in female cancer survivors.

Challenges in the management of Turner syndrome with Y chromosome material: a case report of prophylactic gonadectomy revealing dysgerminoma.

Turner syndrome (TS) patients with Y chromosome material face an increased risk of gonadal germ cell tumors (GCTs). This case report discusses the challenges in decision-making regarding prophylactic gonadectomy, considering the risk of malignancy and the desire to preserve fertility. We report a case of a 12-year-old female with mosaic TS and Y chromosome material who initially presented with short stature and obesity. Karyotype analysis showed a mixed cell line (45X and 46XY). Counseling about the increased risk of developing GCT and preservation of gonadal function was provided, and we decided to delay gonadectomy until the age of 12. Prophylactic bilateral gonadectomy revealed dysgerminoma associated with GB at the age of 12. Fortunately, the patient was asymptomatic, with no additional therapy required due to the early stage of the disease. The case highlights the dilemma in managing TS patients with Y chromosome material, where the risk of GCT varies depending on the type of difference in sex development and gonadal function. The decision to delay gonadectomy reflects the emphasis on preservation of ovarian, although it poses a risk of malignancy. This case underscores the importance of individualized care in TS patients with Y chromosome material, balancing the risk of malignancy against preservation of ovarian. It emphasizes the need for timely and personalized decision-making in prophylactic gonadectomy.

[Polyploidy induction by spherical size standard polystyrene particles in a Chinese hamster cell line CHL].

To investigate relationships between particle (as a model of aggregates) size in a nanomaterial test suspension and its cytotoxicity, a series of eleven sizes of polystyrene (PS) particles were tested in the cytotoxicity test and the chromosome aberration test by using a Chinese hamster cell line CHL. The PS particles were spheres with defined diameters ranging from 0.1 to 9.2 µm. A series of eight sizes of particles with diameters ranging from 0.92 to 4.45 µm showed stronger cytotoxicity than the others. There was a marked difference in cytotoxicity between the 4.45- and 5.26-µm particles. The 0.92- to 4.45-µm particles did not induce structural chromosome aberrations but induced a high frequency of polyploidy in the chromosome aberration test. The 5.26-µm particles showed very weak induction of polyploidy. The incorporation of the 4.45-µm particles into CHL cells was observed by scanning electron microscopy (SEM). Some cells incorporated more than 10 particles. The semi-quantitative measurement of incorporation of particles into cells was performed by flow cytometry with a parameter of side scattered light (SSC) intensity. It showed that CHL cells preferably incorporated the 4.45-µm particles to the 5.26-µm particles. These findings suggest that CHL cells may have a kind of size-recognition ability and incorporate a particular size of particles. The particles may prevent a normal cytokinesis resulting in polyploidy induction. Nanomaterials also may show size-dependent toxicity. Data on particle (or aggregate) size distribution in the test suspension should be provided to evaluate properly the results of toxicity tests of nanomaterials.

Needle fibers of an azo-dye mixture induce polyploidy in a Chinese hamster cell line CHL.

In a routine Safety evaluation of chemicals included in household products, we found a mixture of azo dyes (CMBA, main component: N-[5-[(2-cyanoethyl)ethylamino]-4-methoxy-2-[(5-nitro-2,1-benzisothiazol-3-yl)azo]phenyl] acetamide) that precipitated in the culture medium in a characteristic fiber form (around 2 - 33 microm in length) similar to that of asbestos. We compared CMBA with an asbestos, chrysotile B, in a cytotoxicity, chromosome aberration (CA), and micronucleus (MN) test in a Chinese hamster lung cell line (CHL). In the cytotoxicity test, the 50% growth inhibition concentration was 11.0 microg/ml for CMBA and 0.398 microg/ml for chrysotile B asbestos. CMBA and chrysotile B both induced polyploidy in the CA test and equal-sized binucleated and polynuclear cells in the MN test. CMBA differs from chrysotile B chemically. The former is an organic chemical and the latter is a mineral. Although CMBA is soluble in methanol and can be safely disposed by burning, it should be handled carefully when manufactured in a factory.

Development of an in vitro screening method for safety evaluation of nanomaterials.

To evaluate the role of particle size in cytotoxicity tests of nanomaterials (NMs), we exposed Chinese hamster cells to polystyrene (PS) spheres with defined diameters ranging from 0.1 to 9.2 microm. We found that the 4.45-microm PS particles were most cytotoxic while sizes 0.1 and 0.2 microm showed no cytotoxicity up to 1000 microg/ml. In the chromosome aberration test, the 4.45-microm PS particles induced polyploidy in a mass concentration-dependent manner in 24- and 48-h treatments. The 5.26-microm PS particles induced polyploidy only at 1000 microg/ml for 48 h. Next, we performed the cytotoxicity test with as-grown single walled carbon nanohorns (NHas). These were suspended in DMSO and then transferred into the culture medium followed by sonication. Six suspensions differently sonicated showed the same apparent toxicity, although the total particle size distributions differed. However, the sizes of NHas particles predicted to be most toxic from the experiments with PS particles, i.e. 1.01-4.47 microm constituted 40-60% of all particles in all six suspensions. The results suggest that the cytotoxicity of NMs in suspension depends on specific sizes of aggregates and therefore suspensions should be checked with regard to particle size distributions in assays of toxic effects. The uptake of particles into cells was confirmed by confocal microscopy.

[Change in characteristics of human mesenchymal stem cells during the in vitro culture -- c-myc gene expression and chromosome aberrations at the c-myc locus].

We investigated mRNA expression of c-myc and chromosome aberrations at the c-myc locus in the same passage number of human mesenchymal stem cells (hMSCs). To understand the sensitivity of mRNA expression and the induction of chromosome aberrations, we first tested them in hMSC and cancer cell lines (HeLa S3, HOS, and OUMS-27). The c-myc mRNA expressions in HeLa S3 and OUMS-27 were significantly higher than those in hMSC, but then those in HOS were not. On the other hand, c-myc aberrant cells detected by fluorescence in situ hybridization in HeLa S3, HOS, and OUMS-27 were significantly higher than that in hMSC. Both analyses were performed in hMSCs derived from five donors for the culture period of 50 days. In hMSCs from one donor, the frequency of c-myc aberrant cells significantly increased at 20 and 50 days respectively, and each mRNA expressions had a tendency to increase, but there is no significant change among 3, 20 and 50 days. In hMSCs from the others, both endpoints did not change for 50 days. For safe use of somatic stem cells in the regenerative medicine, the investigation of characteristic change of them during the in vitro culture is important. In the present study, we showed the mRNA expressions and chromosome aberrations of hMSCs in in vitro culture as the first step for establishing of safety evaluation of tissue engineered medical devices using normal hMSCs.

Organic-solvent extraction of model biomaterials for use in the in vitro chromosome aberration test.

We prepared polyurethane (PU) containing 0.4% or 4% 4,4'-methylenedianiline (MDA) as model materials to investigate the effectiveness of sample preparation by organic-solvent extraction for the in vitro chromosome aberration (CA) test. MDA itself (0.4 mg/mL) was positive only in the presence of an exogenous metabolizing system (S9 mix). The culture medium extract of PU containing 4% MDA (PU/4% MDA) was negative with and without S9 mix. Methanol and acetone extracts, on the other hand, induced structural CAs without S9 mix, which we did not expect because MDA requires S9 mix for activity. On chemical analysis, however, we found that the ratio of MDA extracted by the organic solvents to that extracted by the culture medium of PU/4% MDA was about 15:1. Interestingly, oligomers consisting of poly(tetramethyleneglycol) derivatives (OTMG) were also extracted by the organic solvents. The data suggest that the induction of structural CAs in the absence of S9 mix may have been partly due to synergism of MDA and OTMG. CA tests of MDA and PTMG-1000 in combination confirmed that to be the case. Thus, organic-solvent extraction may be more effective than medium extraction in evaluating the biological safety of biomaterials. Detailed chemical analysis of extracts was performed.

[The evaluation of preoperative and postoperative frontal lobe functions in three operative cases of meningioma].

We report three cases of frontal meningioma with their pre- and post-operative evaluations of higher brain functions, especially of frontal lobe functions. All of the cases showed the improvement of the frontal lobe functions after the tumor removal. The evaluations of frontal lobe functions in benign brain tumors such as a meningioma are reported only in a few cases. The evaluations of frontal lobe functions in the operative cases of benign brain tumors provide many interesting and valuable informations about frontal lobe functions. So we must be more interest in evaluations in higher brain functions and accumulate cases for the further analysis of higher brain functions.

The Commelina yellow mottle virus promoter drives companion-cell-specific gene expression in multiple organs of transgenic tobacco.

Previous work has demonstrated that some endogenous plant gene promoters are active in selective companion cells of the phloem, depending on organ types and developmental stages. Here we report that the Commelina yellow mottle virus (CoYMV) promoter is active in the companion cells of leaves, stems and roots of transgenic Nicotiana tabacum cv. Xanthi NN, using beta-glucuronidase (GUS) as a reporter. Thus, the CoYMV promoter has a broad organ specificity. This promoter can be useful in molecular studies on the functions of companion cells in many aspects of phloem biology, such as regulation of long-distance transport, macromolecular traffic, plant development and interaction with pathogens. It may also be useful in engineering crops that produce specific gene products in the companion cells to block long-distance movement of pathogens.

Potato spindle tuber viroid strains of different pathogenicity induces and suppresses expression of common and unique genes in infected tomato.

Viroids are the smallest plant pathogens. These RNAs do not encode proteins and are not encapsidated, and yet they can replicate autonomously, move systemically, and cause diseases in infected plants. Notably, strains of a viroid with subtle differences in nucleotide sequences can cause dramatically different symptoms in infected plants. These features make viroids unique probes to investigate the role of a pathogenic RNA genome in triggering host responses. We conducted a comprehensive analysis of the differential gene expression patterns of tomato plants at various stages of infection by a mild and severe strain of Potato spindle tuber viroid (PSTVd). We also compared tomato gene expression altered by the PSTVd strains with that altered by Tobacco mosaic virus (TMV). Our analyses revealed that the two PSTVd strains altered expression of both common and unique tomato genes. These genes encode products involved in defense/stress response, cell wall structure, chloroplast function, protein metabolism, and other diverse functions. Five genes have unknown functions. Four genes are novel. The expression of some but not all of these genes was also altered by TMV infection. Our results indicate that viroids, although structurally simple, can trigger complex host responses. Further characterization of viroid-altered gene expression in a host plant should help understand viroid pathogenicity and, potentially, the mechanisms of RNA-mediated regulation of plant gene expression.

Plasmodesma-mediated selective protein traffic between "symplasmically isolated" cells probed by a viral movement protein.

Intercellular communication is essential for differentiation and development. In plants, plasmodesmata (PD) form cytoplasmic channels for direct communication. During plant development, programmed reduction in PD number and transport capacity creates the so-called symplasmic domains. Small fluorescent dyes and ions can diffuse among cells within a domain but not across domain boundaries. Such symplasmic isolation is thought to allow groups of cells to differentiate and develop into tissues with distinct structures and functions. Whether or how "symplasmically isolated" cells communicate with one another is poorly understood. One well-documented symplasmic domain is the sieve element-companion cell (SE-CC) complex in the phloem tissue. We report here that, when produced in the CC of transgenic tobacco, the 3a movement protein (3a MP) of Cucumber mosaic virus fused to green fluorescent protein (GFP) can traffic out of the SE-CC complex via PD. The extent of 3a MP:GFP traffic across the boundary between vascular and nonvascular tissues depends on organ type and developmental stage. Our findings provide experimental evidence that endogenous machinery exists for protein traffic between the symplasmically isolated SE-CC complex and neighboring cells. We suggest that PD-mediated traffic of selected macromolecules can be a mechanism for symplasmically isolated cells to communicate with one another.