[Clinical features and gene mutation analysis of patients with Alagille syndrome].

Objective: To analyze the clinical manifestations and gene mutations of patients with Alagille Syndrome (ALGS) to improve diagnosis and provide a boarder spectrum of gene mutagenesis. Methods: A retrospective study was performed in 18 ALGS patients admitted to Xi'an Children's Hospital from January 2016 to January 2020. Clinical characteristics, biochemical parameters, gene mutations and prognosis were collected and analyzed. Next-generation sequencing of liver disease-related gene panels or the whole exome was carried out for the probands. Mutations of candidate genes were verified by Sanger sequencing in their family members. Based on the comparison with a well-known database of disease, the harmfulness and structures of proteins with novel mutations were predicted, and the pathogenicity was evaluated. Results: There were 9 males and 9 females with ALGS in this study, and the age of initial diagnosis was 2.5 (1.9, 6.8) months. All patients initially presented with cholestasis, with other symptoms including 15 cases of special facial features, 11 cases of butterfly vertebrae, 10 cases of congenital heart disease, 5 cases of posterior corneal embryonic ring (among 16 cases with ophthalmological examination), and 1 case of polycystic kidney disease. A total of 14 JAG1 gene mutations and 6 NOTCH2 gene mutations were identified. Among these newly identified mutations, 6 were associated with JAG1 gene, including c.1213delA (p.T405Lfs*7), c.1270dupG(p.A424Gfs*5), c.1741dupG(p.A581Gfs*8), c.3045delC (p.I1016Ffs*20), c.2000-2A>C and c.625C>A(p.H209N); 4 were associated with NOTCH2 gene, including c.6961dupG(p.A2321Gfs*79), c.518G>T(p.G173V), c.6157C>T(p.R2053C) and c.710G>A(p.R237Q). Sixteen patients were followed up for (37.9±31.5) months. Among these cases, 2 died of liver failure (1 case underwent Kasai operation due to misdiagnosis with biliary atresia), 1 improved after liver transplantation, and 13 were in stable condition after medical treatment. Conclusions: The phenotypes of ALGS are diverse, genetic detection can help diagnosis. The JAG1 and NOTCH2 genes showed a wide array of mutations, with many novel mutations identified in this study.

[Pathological significance of NY-ESO-1 expression in the diagnosis of myxoid liposarcoma].

Objective: To detect the expression of New York esophageal squamous cell carcinoma antigen 1 (NY-ESO-1) in common types of mesenchymal myxoid tumors, and to investigate its significance in the diagnosis and differential diagnosis of myxoid liposarcoma. Methods: A total of 43 formalin-fixed paraffin-embedded samples of mesenchymal myxoid tumors from the Affiliated Hospital of Qingdao University and Qingdao Municipal Hospital ranging between 2010 and 2017 were selected. NY-ESO-1 expression was detected by immunohistochemical staining. DDIT3 gene status was detected by fluorescence in situ hybridization (FISH). NY-ESO-1 mRNA was detected by reverse transcription-PCR (RT-PCR). Results: Histopathology and FISH results confirmed that there were 11 cases of myxoid liposarcoma and 32 other types (including 7 cases of well-differentiated liposarcoma, 1 dedifferentiated liposarcoma, 3 lipomas, 2 lipoblastomas and 19 non-adipocytic tumors). Immunohistochemical staining showed that the positive expression propotion of NY-ESO-1 in myxoid liposarcoma was 11/11, and the positive location was the cytoplasm and nucleus of lipoblast cells. The expression intensity is higher in regions with round cell differentiation. Among the 32 cases of other mesenchymal myxoid tumors, only one well-differentiated liposarcoma showed positive immunoreactivity for NY-ESO-1. RT-PCR confirmed that 7 cases of myxoid liposarcoma (7/11) and one well-differentiated liposarcoma (1/7) had NY-ESO-1 mRNA expression. Conclusions: NY-ESO-1 is positively expressed in myxoid liposarcoma. It can be served as a useful marker for the diagnosis and differential diagnosis of myxoid liposarcoma.

EB-virus latent membrane protein 1 potentiates the stemness of nasopharyngeal carcinoma via preferential activation of PI3K/AKT pathway by a positive feedback loop.

Our previous study reported that Epstein-Barr virus(EBV)-encoded latent membrane protein 1 (LMP1) could induce development of CD44(+/High) stem-like cells in nasopharyngeal carcinoma (NPC). However, the molecular mechanisms that underlie modulation of cancer stem cells (CSCs) in NPC remain unclear. Here, we show that LMP1 induced CSC-like properties through promotion of the expression of epithelial-mesenchymal transition-like cellular markers and through alterations in differentiation markers. Furthermore, LMP1 activated and triggered phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) pathway, which subsequently stimulated expression of CSC markers, development of side population and tumor sphere formation. This suggests that PI3K/AKT pathway has an important role in the induction and maintenance of CSC properties in NPC. Similarly, PI3K/AKT pathway was also activated by phosphorylase in LMP1-induced CD44(+/High) cells. In addition, LMP1 greatly increased expression of miR-21 and downregulated expression of the miR-21 target, PTEN. Overexpression of miR-21 by transfection of miR-21 mimics into LMP1-transformed cells led to phosphorylase-mediated activation of the PI3K/AKT pathway and induction of CSCs. On the contrary, phosphorylation of the PI3K/AKT pathway and the expression of CSC were reversed by an miR-21 inhibitor. The specific inhibitor (Ly294002) of PI3K/AKT pathway significantly decreased expression of miR-21 and CSC markers and upregulated the expression of PTEN, which indicates that miR-21 and PTEN are the downstream effectors of PI3K/AKT and that expression of these two effectors are related to the development of NPC CSCs. Taken together, our novel findings indicate that LMP1, PI3K/AKT, miR-21 and PTEN constitute a positive feedback loop and have a key role in LMP1-induced CSCs in NPC.

Unresolved issues in the analysis of F2-isoprostanes, F4-neuroprostanes, isofurans, neurofurans, and F2-dihomo-isoprostanes in body fluids and tissue using gas chromatography/negative-ion chemical-ionization mass spectrometry.

F2-isoprostanes (F2-IsoPs) generated from arachidonic acid (AA) have been recognized as the most reliable marker of nonenzymatic lipid peroxidation in vivo. F2-IsoPs are initially produced in esterified form on phospholipids, and then released into body fluids in free form. The same mechanism can lead to generation of F4-neuroprostanes (F4-NPs) and F2-dihomo-IsoPs from docosahexaenoic acid (DHA) and adrenic acid, respectively. In addition, isofurans (IsoFs) and neurofurans (NFs) may be preferentially produced from AA and DHA, respectively, under high oxygen tension. The detection of F2-IsoPs using gas chromatography/negative-ion chemical-ionization mass spectrometry (GC/NICI-MS) has been widely employed, which is important for human body fluids containing low quantity of free-form F2-IsoPs. F4-NPs have also been detected using GC/NICI-MS, but multiple peaks need to be quantified. In this paper, we summarize the basic workflow of the GC/NICI-MS method for analyzing F2-IsoPs and F4-NPs, and various formats of assays conducted by different groups. We then discuss the feasibility of simultaneous analysis of IsoFs, NFs, and F2-dihomo-IsoPs with F2-IsoPs or F4-NPs. Representative GC chromatograms for analyzing these markers in human body fluids and rat brain tissue are demonstrated. Furthermore, we discuss several factors that may affect the performance of the analysis, such as those related to the sample processing steps, interference from specimens, types of GC liners used, and the addition of electron multiplier voltage in the method setting for the MS detector. Finally, we question the appropriateness of measuring total (free plus esterified) levels of these markers in body fluids.

Serum hormone concentrations and ovarian follicular wave emergence in Jilin sika deer (Cervus nippon hortulorum) after synchronization of estrous cycles.

The study was conducted to investigate the serum hormone concentrations and follicular dynamics present after synchronous treatment (CIDR) in female Jilin sika deer (n = 15) of estrous cycles. Blood samples were collected to analyze the FSH, LH, estradiol and progesterone during the estrous cycles. Manual transrectal ultrasonography examination was conducted at least thrice weekly to monitor the follicular wave. Ultrasonography showed that follicle development occurred in waves, and most estrous cycles in Jilin sika deer consist of one, two, or three waves. The largest follicles of the interwaves of two- and three-wave cycles were different (P < 0.05). The mean interovulatory interval was 15.0 ± 4.6 d. There was a surge in circulating FSH in two- and three-wave cycles. The emergence of the largest follicle was related to the peak of serum concentration of estradiol. Serum progesterone concentrations were not different between one- and three-wave cycles (P < 0.05). We concluded that FSH and estradiol concentration may have an important role in controlling follicular development, that the estrous cycle in Jilin sika deer is characterized by one, two, or three waves of follicular development after synchronization.

Diagnostic value of different patient positions during expiratory low-dose thin-layer MDCT for evaluating air trapping after allogeneic hematopoietic stem cell transplantation.

OBJECTIVE: To investigate the diagnostic value of different patient positions during expiratory low-dose thin-layer multidetector computed tomography (MDCT) for detecting air trapping after allogeneic hematopoietic stem cell transplantation (allo-HSCT). PATIENTS AND METHODS: Expiratory lung MDCT scanning was done for 51 post-allo-HSCT patients in both the supine and prone positions to determine if they had air trapping lesions. We assessed the volume fraction of an air trapping region (CT value of ≤700HU at expiratory phase) against the whole lung area with a GE workstation and graded these results. RESULTS: In the supine position, multiple air trapping lesions were found in 16 of 51 patients, which were scattered and mainly distributed in the dorsal sides of both lower lobes. In the prone position, in addition to these 15 patients, air trapping lesions were also found in 11 other patients, which were mainly distributed in the anterior load-bearing area of the lung lower lobes and lobe-connected areas. Compared with that in the supine position, the graded score of air trapping in the prone position was significantly different (p = 0.006). CONCLUSIONS: When performing expiratory thin-layer MDCT for patients with chronic rejection reactions after allo-HSCT, scanning in the prone position should also be performed, not only to more accurately observe lesions, but also for a preliminary evaluation of air trapping severity. This provides a basis for an early clinical diagnosis and treatment.

Acute myocardial infarction caused by aortic dissection.

Clinical presentation of aortic dissection is similar to that of acute myocardial infarction (AMI). Clinical differential diagnoses from lethal chest pain in emergency department include AMI, aortic dissection, pulmonary embolism, tension pneumothorax, etc. Thrombolytic therapy for recanalization of thrombotic occluded coronary artery in AMI must be considered, but it is absolutely contraindicated for aortic dissection. However, AMI secondary to aortic dissection is a rare condition, which might be caused by compression of the coronary arteries by a hematoma or extension of the dissection into the coronary arterial wall. Surgery is the first choice for AMI secondary to aortic dissection caused by extension of dissection into the coronary arterial wall. We present a case of inferior wall AMI caused by type I aortic dissection with presentation of chest pain and hemiparaplegia of right lower limb.

Mutation analysis of the putative tumor suppressor gene PTEN/MMAC1 in cervical cancer.

OBJECTIVE: PTEN/MMAC1, a candidate tumor suppressor gene located at chromosome 10q23.3, was recently identified and found to be homozygously deleted or mutated in several different types of human tumors. The aim of this study is to determine whether PTEN/MMAC1 is a target for 10q loss of heterozygosity in cervical cancer. METHOD: We examined 50 primary cervical carcinoma specimens using a PCR-based assay followed by SSCP and direct sequencing. The genomic DNA was also confirmed by Southern blot analysis. RESULTS: All specimens except one, which has a 7-base deletion, showed a negative result. Among them, 30 randomly selected cases and their paired noncancerous tissue were further screened using nested RT-PCR. Six of 30 cervical cancerous tissues had aberrant transcripts. However, 4 of the matched noncancerous tissues also had aberrant transcripts. Southern blot analysis of the entire genomic DNA did not reveal any evidence of gene alteration. CONCLUSIONS: Sequence abnormalities in the PTEN/MMAC1 gene were only detected in 1 of 50 cervical cancers analyzed indicating that aberrant PTEN/MMAC1 function is an uncommon event in the development of cervix cancers. However, similar to studies with the TSG101 gene, screening for aberrant transcripts of PTEN/MMAC1 with nested RT-PCR may detect transcripts, which, although they vary from the normal size, may not be related to oncogenesis as they are also frequently found in normal tissues of the same patient.

Is modified in situ vaginal wall sling operation the treatment of choice for recurrent genuine stress incontinence?

PURPOSE: We evaluate objectively the results of a modified in situ vaginal wall sling operation for recurrent genuine stress incontinence and whether it is a substitute for the traditional sling procedure. MATERIALS AND METHODS: A total of 23 patients with urodynamically proved recurrent genuine stress urinary incontinence were recruited in this study. Patients were treated with a modified needle urethropexy technique using an island of in situ vaginal skin as a sling to support the bladder neck and urethra. Surgical outcome was evaluated subjectively and objectively at a median of 15 months. A total of 42 patients who underwent a traditional polytetrafluoroethylene sling operation served as controls. RESULTS: The cure rate of the vaginal wall sling operation was 34.8% by objective assessment, which was lower than that of the traditional sling procedure (88.1%, p <0.05). The subjective success rate demonstrated the same results (vaginal sling 60.9% versus traditional sling 92.9%, p <0.05). The risk factors for operation failure were lower maximal urethral pressure, lower urethral closing pressure, narrow vaginal capacity and previous anterior colporrhaphy or a Stamey operation (all p <0.05). In 3 cases suburethral epithelial inclusion cysts were specific complications of the operation. There was no prolonged urinary retention or urethral erosion. CONCLUSIONS: Based on our results, we do not believe that the vaginal wall sling operation should be recommended for all recurrent genuine stress urinary incontinence cases and especially not for those with factors predictive of surgical failure. Further studies are needed to investigate and clarify the possible causes of failure.

Analysis of TSG101 tumour susceptibility gene transcripts in cervical and endometrial cancers.

Carcinoma of the uterine cervix is a common malignancy among women that has been found to show loss of heterozygosity in the chromosome 11p. Recent studies have localized the TSG101 gene in this region, and also demonstrated a high frequency of abnormalities of this gene in human breast cancer. To determine the role of the TSG101 gene in the carcinogenesis of cervical and uterine carcinoma, 19 cases of cervical carcinoma and five cases of endometrial carcinoma, as well as nearby non-cancerous tissue from the same patients, and 16 blood samples from healthy persons as normal control were analysed by Southern blot analysis of genomic DNA, reverse transcription of the TSG101 mRNA followed by PCR amplification and sequencing of the products. We found that abnormal transcripts of the TSG101 gene were common both in cancerous or non-cancerous tissues of the uterus and cervix and in normal peripheral mononuclear cells. There was no genomic deletion or rearrangement in spite of the presence of abnormal transcripts, and no definite relationship between the abnormal transcripts and HPV infection was found. Although the frequency of abnormal transcripts was higher in cancerous than in non-cancerous tissue, normal peripheral mononuclear cells also had abnormal transcripts. Given these findings, the role of the TSG101 gene as a tumour-suppressor gene should be re-evaluated. Because some aberrant transcripts could be found at the first PCR reaction, we suggest that the aberrant transcripts might be the result of imperfect minor splicesome products.

Analysis of microsatellite instability in cervical cancer.

Microsatellite instability was first reported in hereditary nonpolyposis colorectal cancer (HNPCC) as well as other cancers, including endometrial and ovarian cancers. Single base repeat markers of human MSH3 and MSH6 genes were found to precipitate the action of human MSH2. The marker BAT-26 was reported to be a simple, low-cost, and rapid marker for detection replication errors (RER) and the status of colorectal cancers. We analyzed di-nucleotide repeats of the microsatellite markers (D2 S123, D5 S82, D5S299, D10S197, D17S791, D18S34), single base repeat markers (DeltaP3, hMSH3, hMSH6, and TGFbeta-RII), and BAT-26 to evaluate microsatellite instability in cervical cancer. Altogether 80 paired cervical cancers were studied. Our results showed that microsatellite instability is not common in cervical cancer, and the mutation of the single base repeat of mismatch repair (MMR) genes (hMSH3 and hMSH6) is also uncommon. The BAT-26 is not a good marker to detect the RER status of cervical cancer.

Analysis of FHIT transcripts in cervical and endometrial cancers.

Carcinoma of the uterine cervix is a common malignancy, and many affected women, have been found to exhibit loss of heterozygosity (LOH) in the chromosome 3p region. Recent studies have localized the FHIT (fragile histidine triad) gene in this region and also demonstrated a high frequency of abnormalities of this gene in various cancers. To determine the role of the FHIT gene in cervical and uterine carcinomas, 16 cases of cervical carcinoma and 7 cases of endometrial carcinoma, as well as nearby non-cancerous tissues in these patients, were analyzed by reverse transcription of the FHIT mRNA followed by polymerase chain reaction amplification and sequencing of the products. In this study, 13 of 16 cervical cancers and 4 of 7 endometrial cancers displayed abnormal FHIT transcripts, including a lack of 2 or more exons of the FHIT gene, the insertion of several bases in the deletion junctions, and a 282 bp deletion from cDNA 171 to 452, resulting in a frameshift. Moreover, 5 of 16 matched non-cancerous tissues from the cervical cancer patients and 4 of 7 non-cancerous tissues from endometrial cancer patients also showed the presence of abnormal transcripts lacking 3 or more exons of the FHIT gene. Only 1 of 23 paired samples exhibited LOH. Our results suggest that the abnormal transcript of the FHIT gene is common in both normal and tumor tissues of the uterus and cervix. We also checked for HPV infection in these samples and found no definite relationship between the abnormal transcript and human papillomavirus infection.

Prospective comparison of laparoscopic and traditional colposuspensions in the treatment of genuine stress incontinence.

OBJECTIVE: To compare prospectively the results of laparoscopic and traditional colposuspensions in the treatment of genuine stress incontinence and to evaluate the efficacy, technique, and functional and anatomical changes after these two procedures. MATERIALS AND METHODS: Ninety-two patients with urodynamically proven genuine stress incontinence participated in this study, with 46 patients randomly allocated to laparoscopic colposuspension, and the other 46 patients to the traditional procedures. All patients had repeat studies at least 3 months after operation. RESULTS: The bladder neck position was significantly elevated after operation either at rest or during straining in both groups (all p < 0.001), but it was higher in the traditional group than the laparoscopy group during straining (p < 0.05). Comparison of urodynamics before and after operation in both groups showed significantly increased minimal urethral resistance and improved pressure transmission ratios at the proximal urethra (Q2). The blood loss was less in the laparoscopy group. The duration of bladder drainage after laparoscopic colposuspension was shorter, and was not affected by subsequent laparotomy. The operative time was almost the same. The success rate of the laparoscopy group was lower than that of the traditional group (80.4% vs. 95.6%, p = 0.044). The complication rates were 10.8% and 17.4% respectively. CONCLUSION: Laparoscopic colposuspension is an effective method for the treatment of GSI, as documented by anatomical and functional assessments. However, the success rate is still lower than for the traditional procedure.

Mini-catheter used for bladder drainage following stress-incontinence surgery and the factors relating to drainage failure.

OBJECTIVES: To determine the most effective length for the mini-catheter and to discuss factors relating to drainage failure. METHODS: Prospective study of a No. 8 silastic feeding tube for bladder drainage omits the necessity of additional catheterization for measuring residual urine. This two-part study in which 146 patients took part, first, determined the suitable length for insertions for the perforated portion and the imperforated portion; then, gauged the effectiveness of the mini-catheter with desired proportions. RESULTS: Drainage is better when the imperforated portion is at least 5 cm in length (p = 0.0129). The length of the perforated portion, on the other hand, has less impact of the resultant drainage provided that its length does not exceed 5 cm. Beyond this length, an obstruction often occurs. There were 2 obstructions (1.3%) induced by blood clot as well as 9 patients (5.8%) who suffered from one, or more, complications attributed to the mini-catheter for reasons other than the length of insertion. CONCLUSIONS: When care is taken in creating a mini-catheter of the proper proportions, it offers many advantages and very few complications to gynecologists and patients alike.

Specific overexpression of P53 in sarcoma derived from Rat-1 cells transformed by cigarette smoking condensate-treated human fetal lung DNA.

The p53 protein is highly expressed in fibrosarcoma derived from Rat-1 cells transfected with cigarette smoke condensate-treated human fetal lung DNA but expressed low in the counterparts of nitroso-N-methylurea and dimethyl sulfoxide groups. Our results denoted that a high expression of p53 protein specifically contributes to the initiation of human lung carcinogenesis induced by cigarette smoke condensate. High expression and missense mutation of p53 may probably be a potential biomarker of initiation of human lung carcinogenesis.

Inhibition of the transforming ability of cigarette smoking condensate-treated human fetal lung DNA induced by oltipraz.

Rat-1 cells transfected by genomic DNA of human fetal lung explants treated with 100 micrograms/ml of oltipraz (5-(2-pyrazimyl)-4-methyl-1, 2-dithiol-3-thione) for 14 hr or 100 micrograms/ml of cigarette smoking condensate for 6 hr formed 0 to 8 transformation foci, respectively. If 100 micrograms/ml of oltipraz was added to culture of human fetal lung explants 8 hr prior to the treatment of cigarette smoking condensate, the Rat-1 cells transfected by genomic DNA of human fetal lung explants formed only two foci. In addition, the growth speed of Rat-1 cells transfected by genomic DNA of human fetal lung treated with both oltipraz and cigarette smoking condensate was lower than that transfected by cigarette smoking condensate-treated human fetal lung DNA. Our results indicate that oltipraz can block the irreversible change of human fetal lung DNA caused by cigarette smoking condensate, and the results suggest the possibility of using oltipraz as control in the experimental initiation of human lung carcinogenesis.

[Influence of selenium supplement on cadmium metabolism in human].

38 subjects were randomly divided into 2 groups of 19 each. Group 1 consumed a selenium supplement (150 micrograms/d X 21) and Group 2 received only placebo(glucose). After supplementation, blood Se levels and plasma GSH-Px activities in Group 1 increased from 76 to 100 ng/ml (P less than 0.05) and 0.082 to 0.122 e.u./ml (P less than 0.01) respectively. All measured Se, GSH-Px values in Group 2, and high concentrations of lipid peroxides (greater than 4 nmol/ml as malonaldehyde) in both groups remained approximately the same. Se supplementation resulted in a marked decrease of RBC cadmium (Cd) from 32.3 to 25.4 micrograms/g Hb (P less than 0.001). Urinary and fecal Cd in 5 subjects of each group were analyzed every 4 days, and the results demonstrated that Cd was mainly excreted in feces after Se supplementation. One week after discontinuing of Se treatment, Cd content in urine and feces decreased to control levels. Theoretical evidence for chemoprevention of lung cancer with Se in this area was thus provided.

[Effect of molybdenum and tungsten on mammary carcinogenesis in Sprague-Dawley (SD) rats].

Virgin female rats of SD strain were given ad libitum a nutritionally adequate semipurified diet containing 0.026 ppm molybdenum and deionized water (groups 1-3) or the same diet with 150 ppm tungsten and the drinking water (group 4). Group 1 was used as control. After 15 days, all the animals in groups 2-4 received an intravenous injection of N-nitroso-N-methylurea (NMU) 5 mg/100 g body weight. One week after administration of carcinogen, 10 ppm Mo was added to the drinking water in group 3. After 125 days, the mammary cancer incidence in group 4 (79.2%) was significantly higher than that in group 2 (50.0%) or group 3 (45.5%) (P less than 0.05). After 198 days, the average number of mammary cancer in each animal and mammary cancer incidence in group 3 (1.5 and 50.0%) were obviously lower than those in group 2 (2.0 and 90.5%) or group 4 (2.6 and 95.7%). The first palpable mammary tumor was found in the W-supplemented group only 56 days after the injection of NMU, whereas in the W-unsupplemented and Mo-supplemented groups, the first mammary tumor was observed 71 and 85 days after NMU treatment. Of these 181 mammary tumors, 177 (97.8%) were adenocarcinoma or papillary carcinoma, only 4 (2.2%) fibroadenocarcinoma. The results of this study show, for the first time, the inhibitory effect of Mo on the mammary carcinogenesis and promoting effect of Tungsten, an antagonist of molybdenum, on the tumor growth.

Effects of molybdenum and tungsten on mammary carcinogenesis in SD rats.

Virgin female rats of the SD strain were fed ad libitum a nutritionally adequate semipurified diet and demineralized water (groups 1 and 2), or the same diet with 10 ppm molybdenum (group 3) or 150 ppm tungsten (group 4) added to the drinking water. The animals in groups 2-4 received a single iv injection of 5 mg N-nitroso-N-methylurea (NMU; CAS: 684-93-5)/100 g body weight at 50 days of age. One hundred and twenty-five days after the NMU treatment, group 2 exhibited a 50.0% incidence of mammary carcinoma. Group 4 exhibited a significant increase in carcinoma incidence (79.2%) and the value for group 3 (45.5%) was not significantly different from that of group 2. The carcinoma incidence of group 3 (50.0%) was significantly lower than that of group 2 (90.5%) or group 4 (95.7%) 198 days after NMU treatment.