Yoshio Hiasa

Immunohistochemical Analysis of Midkine Expression in Human Prostate Carcinoma

Midkine (MK) is a growth/differentiation factor frequently expressed at high levels in some types of human malignancies. To investigate whether MK is a useful marker in prostate carcinogenesis, immunohistochemical analysis was performed on samples of both latent and clinical prostate cancers of various stages, as well as on specimens of normal gland and prostatic intraepithelial neoplasia (PIN). Of the 80 clinical cancers examined, 69 specimens (86.3%) were immunoreactive for MK, with metastatic lesions generally showing higher expression than the corresponding primaries; normal prostate tissues were negative or showed only weak staining. Midkine was also detected in 12 of 15 latent cancers (80%) and in 12 of 16 cases of PIN (75%). In sections of whole prostate, MK showed variable expression through tumorous sections, probably in reflection of heterogeneous cell populations. The results demonstrate the possible value of MK as a marker for early and latent disease, as well as for more advanced clinical stages of prostate cancer.

Hypertonic saline resuscitation reduces apoptosis and tissue damage of the small intestine in a mouse model of hemorrhagic shock.

&NA; The effect of hypertonic saline resuscitation on intestinal damage and the incidence of apoptosis after hemorrhagic shock were investigated. After anesthesia, male BALB/c mice weighing 24‐34 g were hemorrhaged to the mean arterial pressure of 40 ± 5 mmHg for 90 min. Animals were randomly assigned to four groups: 1) resuscitation with 4 mL/kg of 7.5% NaCl (hypertonic saline; HS) + shed blood (SB); 2) resuscitation with two times the volume of shed blood of lactated Ringers solution (2LR) + SB; 3) sham (catheter only); or 4) control (no treatment). Intestinal damage was graded based on the extent of the vacuolation at the basal area of the intestinal villi. Apoptosis of the small intestines was examined with the terminal deoxynucleotidyl transferase‐mediated deoxyuridine 5‐triphosphate nick‐end labeling method and with DNA laddering. Caspase‐3 activation, heat shock protein (HSP) 70, and HSP40 were assessed by western blotting. Apoptosis of the small intestine and intestinal damage were significantly lower (P < 0.01) in the HS+SB group compared with the 2LR+SB group 2 h and 6 h after hemorrhagic shock and resuscitation, respectively. This corresponded with more DNA fragmentation in the small intestine of the 2LR+SB group compared with the HS+SB group 2 h after hemorrhage and resuscitation. In addition, we observed less caspase‐3 activation in the small intestine of the HS+SB group compared with the 2LR+SB group at 2 h after resuscitation. The content of HSP40 and HSP70 in the HS+SB group was similar to that in controls, but slightly decreased in the 2LR+SB group. HS resuscitation reduced intestinal damage and apoptosis after hemorrhagic shock, suggesting that HS resuscitation may improve the outcome after hemorrhagic shock by reducing apoptosis and damage to the small intestine.

Comparison of ras activation in prostate carcinoma in Japanese and American men.

Comparative studies of point mutations in K‐, N‐, and H‐ras oncogenes were performed on prostate carcinoma from Japanese and American patients to clarify the racial difference.

Clinical studies of de novo aneurysms.

Although multiple cerebral aneurysms ae well recognized, a new aneurysm has only rarely been documented after successful treatment for an aneurysm elsewhere. In our consecutive series of 986 patients with intracranial saccular arterial aneurysm collected from 1975 to 1990, nine patients who had previously unverified (hence, de novo) intracranial aneurysms and ruptures at intervals of 4 to 7.5 years after clipping of an initial aneurysm are presented here. All nine had undergone successful treatment of a previous aneurysm; preoperative and postoperative angiography showed not only successful clipping of the first aneurysm but also no incidence of multiple aneurysms. These patients had suffered from hypertension before their second admission. Seven of the nine patients were treated surgically. All patients had experienced angiographical or symptomatic vasospasm after the first subarachnoid hemorrhage. In the second admission however, seven patients who underwent the surgery for a new aneurysm suffered from no vasospasm in spite of the prominent second subarachnoid hemorrhage. Two of the nine patients died of primary brain damage due to the hemorrhage and underwent necropsy. A histological study of a new aneurysm demonstrated the same findings as that of a usual saccular aneurysm. This clinical study of our patients suggests that it is important to control blood pressure for protection against a new aneurysm formation.

Immunohistochemical Detection of p53 Oncoprotein in Human Oral Squamous Cell Carcinomas and Leukoplakias: Comparison with Proliferating Cell Nuclear Antigen Staining and Correlation with Clinicopathological Findings

Forty oral squamous cell carcinomas and 20 leukoplakias were examined for expression of p53 oncoprotein using an immunohistochemical technique with BP53-12 monoclonal antibody. Positive staining was found in 21/40 (52%) of the carcinomas and 2/20 (10%) of the leukoplakias. Furthermore, comparison of p53 expression with binding of PC10 monoclonal antibody to proliferating cell nuclear antigen (PCNA) and degree of histological malignancy in terms of invasion and histological differentiation of carcinomas demonstrated a positive correlation in both cases.

Expression of the estrogen receptor in human thyroid neoplasms

The expression and quantitation of the estrogen receptor (ER) in human thyroid tumors were examined by biochemical, immunohistochemical, and reverse transcriptase-polymerase chain reaction (RT-PCR) techniques. For this study, neoplasms, adenomatous goiters and adjacent normal thyroid tissues were obtained from 35 patients which included 10 cases of papillary carcinomas, 17 cases of adenomas and 8 cases of adenomatous goiters. Regardless of the histopathological subtype, ER was detected in 19% (5/27) of the neoplastic tissues with the mean value of ER content of 5.0 +/- 1.3 fmol/mg protein and the mean Kd value of 0.38 +/- 0.28 nM. ER was also detected, but at a lower concentration (2.8 +/- 1.6 fmol/mg protein), in the surrounding normal tissues. There was no significant difference between the neoplasms and adenomatous goiters with respect to the incidence of ER positivity and ER content. Furthermore, ER-positive specimens, as determined by both biochemical and immunohistochemical techniques, also showed the expression of ER mRNA detected by RT-PCR method. These results demonstrate that both ER mRNA as well as ER protein are expressed in thyroid neoplasms. This suggests the possibility that estrogen may affect the tumorigenesis or the progression of some thyroid neoplasms.

Immunohistochemical analysis of estrogen receptors in 313 paraffin section cases of human thyroid tissue

Three hundred and thirteen cases of human thyroid tissues, comprising 39 nodular goiters from 34 females and 5 males, 130 adenomas from 93 females and 37 males, and 144 carcinomas from 99 females and 45 males were used for the present immunohistochemical assessment of estrogen receptor (ER) expression. Thirty-three cases of follicular carcinoma, 115 cases of papillary carcinoma and 6 cases of anaplastic carcinoma were included in the malignant tumor group. Incidences of ER-positive cases were 23/39 (58.9%) for nodular goiter, 44/130 (33.8%) for adenoma and 26/144 (18.0%) for cancer. In the individual carcinoma categories, 7/23 (30.4%) follicular, 19/115 (16.5%) papillary and 0/6 (0%) anaplastic lesions were judged as positive cases. Thus, the incidence of ER-positive cases tended to decrease with the degree of malignancy; this trend being similar in both sexes. Moreover, the average ages of ER-positive cases were lower than those of ER-negative cases for all types of thyroid carcinoma except the follicular variety in males. It was thus suggested that ER expression may be related to prognosis and tumor growth at early stage. Since the incidence of ER does not significantly differ between females and males, the observed sex differences regarding thyroid tumor incidence may reflect the higher estrogen serum content in females.

Immunohistochemical evaluation of estrogen receptor status in benign prostatic hypertrophy and in prostate carcinoma and the relationship to efficacy of endocrine therapy.

The levels of estrogen receptors in human benign prostatic hypertrophy and in various pathological classifications of prostate carcinoma were assessed using immunohistochemical methods. All cases of benign hypertrophy showed elevated levels of estrogen receptor, while receptor-positive cells were detected in only 48% of carcinomas, indicating a negative correlation between receptor status and malignancy. Furthermore, the prognosis for effective endocrine therapy was poor in cases where tissues demonstrated low or negative receptor levels. In addition, the estrogen receptor status was compared to cell kinetic index such as proliferating cell nuclear antigen and argyrophilic staining of the nuclear organizer region.

Experimental Induction of Renal Tumors

Renal tumors have been reported to be induced by natural products, chemical carcinogens, viruses, or radiation. Species or strain specificity and sex also appear to play significant roles in their development. In man, it is also likely that the heredity existence of other diseases, smoking, food habits, and irradiation may be etiological factors. It also appears that hormonal, chemical, and other environmental factors can play a role. Nephrotoxin modifies two-stage chemical carcinogenesis in rat kidney. Some nephrotoxins without carcinogenicity promote the development of renal tumors in rats pretreated with subcarcinogenic doses of chemical carcinogens. The importance of nephrotoxin in development of renal adenocarcinomas needs elucidation. Preneoplastic lesions in the kidneys can be recognized by histochemical methods with specific antibodies. It is hoped that further research will be continued, so that data obtained from experimental work will provide a better understanding of the etiology and induction of renal cancer in man.

Activation of the MN/CA9 gene is associated with hypomethylation in human renal cell carcinoma cell lines

The MN/CA9 (G250) gene expressed in the normal alimentary tract in a tissue‐specific manner is often activated in renal cell carcinomas. To cast light on the activation mechanism, we examined the methylation status of this gene in seven human renal cell carcinoma cell lines (SKRC‐01, ‐06, ‐10, ‐12, ‐14, ‐44, and ‐59) and three normal kidney tissue samples by using the bisulfite genomic sequencing protocol. CpG methylation was measured at seven locations in the MN/CA9 5′ region. MN/CA9 transcripts were detected by reverse transcription–polymerase chain reaction in five of the renal cell carcinoma cell lines (SKRC‐01, ‐06, ‐10, ‐44, and ‐59). These MN/CA9 positive cell lines showed hypomethylation, whereas the remaining two cell lines (SKRC‐12, and ‐14), and three normal kidney tissue samples without transcripts demonstrated hypermethylation. Treatment with the demethylating agent 5‐aza‐2′‐deoxycytidine resulted in activation of the MN/CA9 gene in the negative cell lines (SKRC‐12 and ‐14). These data suggest that hypomethylation in the 5′ region may have a major role in expression of the MN/CA9 gene in renal cell carcinoma cells. Mol. Carcinog. 27:184–189, 2000.