Determining the rotational alignment of the tibial component at total knee replacement: a comparison of two techniques.

We prospectively assessed the benefits of using either a range-of-movement technique or an anatomical landmark method to determine the rotational alignment of the tibial component during total knee replacement. We analysed the cut proximal tibia intraoperatively, determining anteroposterior axes by the range-of-movement technique and comparing them with the anatomical anteroposterior axis. We found that the range-of-movement technique tended to leave the tibial component more internally rotated than when anatomical landmarks were used. In addition, it gave widely variable results (mean 7.5 degrees ; 2 degrees to 17 degrees ), determined to some extent by which posterior reference point was used. Because of the wide variability and the possibilities for error, we consider that it is inappropriate to use the range-of-movement technique as the sole method of determining alignment of the tibial component during total knee replacement.

Relationships between matrix metalloproteinases and tissue inhibitor of metalloproteinases in the wall of abdominal aortic aneurysms.

AIM: The pathologic feature of aortic aneurysm is considered to be the remodeling of the aortic wall, involving fragmentation and decrease of elastic fibers in the tunica media. Matrix metalloproteinases (MMPs), particularly MMP-2 and MMP-9, have been implicated in collagen and elastin degeneration within the aortic wall. The precise relationship among MMPs and tissue inhibitor of metalloproteinases (TIMPs) is still unclear. We have studied the expression of MMP-2, MMP-9 tissue inhibitor of metalloprotein-1 (TIMP-1), TIMP-2 and membrane type 1-MMP (MT-1-MMP) in the wall of small AAAs (30-45 mm), large AAAs (>45 mm) and controls (<25 mm). We investigated the relationship among expressions of MMP-2, TIMP-2 and MT1-MMP in the walls. METHODS: The aortic walls in the patients with AAA were harvested from the maximum diameter, while the aortic walls in autopsy cases were harvested as controls. We analyzed tissue distribution of cell types by immunochemistry, protein expression by Western blotting and mRNA expression by competitive polymerase chain reaction. RESULTS: They consisted of 11 in controls, 8 in small AAAs and 26 in large AAAs. Among the MMPs-positive-cells, mainly macrophage, MMP-2-positive cells were in the intima, but MMP-9-positive cells in the intima and adventitia. In the small size, MMP-2 and MMP-9 mRNA were higher than those of control. In the large size, MT1-MMP and MMP-9 mRNA were higher than those of the controls. In the mRNA level of the whole AAA, significant correlations were present between MMP-2 and MMP-9, between MMP-2 and TIMP-1, and between MMP-9 and TIMP-1. These expressions were confirmed by Western blotting. CONCLUSION: We concluded as follows: 1) MMP-2 and MMP-9 may play an important role in the developmental process of AAA. 2) TIMP-1 plays an important role of interacting MMP-2 and/or MMP-9. 3) MMP-2 and MT1-MMP may play an important role in the early stages of AAAs.

Paradoxical enhancement of spinal-cord-evoked potentials rostral and caudal to the site of progressive cord compression in the cat.

STUDY DESIGN: Analysis of the sequential waveform changes of the spinal-cord-evoked potentials (SCEPs) associated with progressive cord compression in the cat. OBJECTIVES: To document the phenomenon of paradoxical enhancement of SCEPs despite conduction abnormalities and to evaluate its possible significance. SETTING: Kochi Medical School, Kochi, Japan. METHODS: SCEPs were recorded simultaneously at four serial intervertebral levels, from T6-7 to T9-10 caudal to, and at three serial levels from T2-3 to T4-5 rostral to the compression site at T5-6 following epidural stimulation at L6 in 14 cats. RESULTS: Caudal to the compression site, the area of negative peak significantly increased toward maximal values of 277+/-36 (mean+/-SE), 151+/-9 and 110+/-4% as compared to the baseline precompression values (100%) at T6-7, T7-8, and T8-9, respectively. Rostral to the compression site, the area of negative peak significantly increased before subsequent deterioration and reached 105+/-2, 106+/-2, and 104+/-2% at T4-5, T3-4, and T2-3, respectively. The onset of negative peak enhancement, recorded either caudal or rostral to the compression site, showed a close temporal correlation (r>0.8, P&<0.001) with that of the prolongation in latency of SCEPs at T2-3. CONCLUSIONS: A progressive focal conduction block induced by compression of the spinal cord can paradoxically enhance the ascending SCEPs both caudally and, though less consistently, rostrally, representing a warning of the impending risk of paraplegia.

Effect of lactational exposure to 1,2,3,4- tetrachlorodibenzo-p-dioxin on cytochrome P-450 1A1 mRNA in the neonatal rat liver: quantitative analysis by the competitive RT-PCR method.

BACKGROUND AND METHODS: The aim of this study was to assess the effect of lactational exposure to dioxins in neonates on the cytochrome P450 1A1 (CYP1A1) induction in the level of gene expression. Maternal rats were treated with a single dose of 50 or 100 micromol/kg 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-TCDD), a low potent congener of dioxins, on the first day post-partum (day 1). Induction of CYP1A1 mRNA expression was quantitatively analyzed by the competitive reverse transcription-polymerase chain reaction (RT-PCR) method. RESULTS: The CYP1A1 mRNA was detectable at extremely low amounts in the liver of control neonates and mothers. The mRNA ratios of CYP1A1 to beta-actin in neonates were dose-dependently increased by the treatment of 1,2,3,4-TCDD of their mothers. Its peak occurred on day 6 and was sustained at the same level on day 10. Increases of the ratio with 100 micromol/kg 1,2,3,4-TCDD on day 2, 6 and 10 were 26-, 40- and 40-fold of the appropriate controls, respectively. These levels paralleled the activity of ethoxyresorufin-o-deethylase, representing CYP1A mediated monooxygenase. In the mother, the mRNA ratio was increased only to threefold of the control, 10 days after treatment. CONCLUSION: Current RT-PCR procedure enabled to assess both constitutive and induced levels of CYP1A1 mRNA in the neonatal rat livers. Although the dose of 1,2,3,4-TCDD selected in this study was about 5000 times higher than the daily intake of dioxins in breast-fed infants, CYP1A1 mRNA was highly induced for a longer period of time in neonatal rats receiving 1,2,3,4-TCDD via lactation than the treated maternal rats.

Mn-SOD antisense upregulates in vivo apoptosis of squamous cell carcinoma cells by anticancer drugs and gamma-rays regulating expression of the BCL-2 family proteins, COX-2 and p21.

ROIs and their scavengers are associated with apoptosis induction by anticancer drugs and gamma-rays, but the details have not been clarified. We examined the effect of transfection of Mn-SOD antisense on apoptosis by 5-FU, PLM, CDDP and gamma-rays using nu/nu mice. After inoculation of Mn-SOD antisense-transfected SCC cells into the subcutis of each mouse's back, they slowly multiplied to form tumors sized 1,460 +/- 70 mm(3) at day 60, while control vector-transfected SCC cells rapidly multiplied, with a mean tumor size of 2,330 +/- 220 mm(3). Inversely, mice in the Mn-SOD antisense group survived longer (mean survival duration 94.4 +/- 12.7 days) compared to those in the empty vector group (67.3 +/- 6.8 days). After treatment with 5-FU (5 microg/day), PLM (50 microg/day), CDDP (10 microg/day) and gamma-rays (2 Gy/day), mean survival times were largely prolonged, to 126.3 +/- 22.7, 123.0 +/- 22.1, 136.3 +/- 24.0 and 143.0 +/- 20.8 days, respectively, while mean survival times in the empty vector group were 91.7 +/- 14.8, 85.7 +/- 13.3, 97.5 +/- 16.0 and 100.7 +/- 17.1 days, respectively. Immunohistologically, tumors in the Mn-SOD antisense group revealed additional nick end-labeled cells compared to those in the empty vector group. In comparison, strong expression of Bax, Bak and p21(waf1/cip1) and suppressed expression of Bcl-2, Bcl-X(L) and COX-2 were observed in the Mn-SOD antisense group and the expression pattern of these proteins was the inverse in the empty vector group. The increased expression of these proapoptotic proteins appeared to be p53-independent because p53 protein expression was not increased in the antisense group. These immunohistologic results were supported by Western blotting of each protein. In conclusion, Mn-SOD antisense transfection is advantageous for apoptosis induction of SCC cells by anticancer drugs and gamma-rays through induction of proapoptotic Bcl-2 family proteins and suppression of antiapoptotic protein expression.

[Influence of peplomycin on pulmonary function (PaO2, %DLco) in patients with oral carcinoma].

The pathogenesis of pulmonary fibrosis (PF) induced by bleomycin and its derivative, peplomycin (PEP), is insufficiently understood. To prevent PF and to administer PEP safely, we examined the influence of PEP on pulmonary function in 135 patients who underwent concomitant chemo (PEP + 5-FU)-radio (60Co) therapy and pulmonary function tests. In the inductive therapy, 5 mg of PEP was intramuscularly injected three times a week and a total of 41.6 +/- 14.3 mg was administered. Of the patients, 98 received oral azelastine hydrochloride (AZH, 4 mg/day) during the inductive therapy with the aim of prophylaxis of PF. The oxygen partial pressure in arterial blood (PaO2) only slightly decreased from 84.2 +/- 12.1 mmHg before treatment to 82.8 +/- 12.5 mmHg after treatment, while, carbon oxide diffusion (%DLco) decreased after treatment in most patients (p < 0.001, by paired t test) with mean values before treatment of 106.3 +/- 24.5% and after treatment 99.5 +/- 24.9%. The decrease of %DLco was associated with the dose of PEP until about 40 mg but further decreases of %DLco were not prominent. In the patients who underwent oral AZH, the decrease of %DLco weaker than that in patients without AZH: the decrease rates of %DLco in the former and latter were 4.3 +/- 9.4% and 14.1 +/- 15.9%, respectively. From the chest X-ray examination, mild PF was suspected in three patients but no advancement of PF or clinical symptoms were observed. From these results, it was concluded first that %DLco is more useful than PaO2 as the predisposing risk factor for PF, second that the decrease of %DLco depends on the dose of PEP until about 40 mg, third that AZH is expected to inhibit PEP-induced PF, and fourth that a small dose (20-40 mg) of PEP can be administered without inducing PF if care is exercised as to the patient's age, general condition and the value of %DLco in the use of PEP.

The upregulation by peplomycin of signal transduction in human cells.

To explore the mechanism of pulmonary fibrosis by bleomycin and its derivative, peplomycin (PLM), we examined the influence of PLM on signal transduction in human peripheral blood lymphocytes (HL), monocytes (HM) and fibroblasts (HF). Tyrosine phosphorylation of multiple proteins in HL and HM were induced by 0.001 to 0.05 microg/ml and by 0.01 to 0.5 microg/ml of PLM, respectively. In HF, 116-kDa protein was phosphorylated 0.2 to 5 microg/ml of PLM. When HL were treated with 0.01 microg/ml of PLM, phosphorylation of p56lck and activation of extracellular-signal related kinase-2 (ERK2) were induced. ERK2 was also activated in HM. Coordinately, the ratio of p21ras-binding GTP/GDP was increased by PLM. As well as interleukin-2, PLM induced tyrosine phosphorylation of JAK-3. In addition, PLM upregulated the nuclear translocation of nuclear factor-kappa B and the expression of c-myc-mRNA in HL, HM and HF. Furthermore, 0.01 to 0.001 microg/ml PLM enhanced the cytokine generation by HL and HM, and 1 to 5 microg/ml PLM increased cytokine generation and collagen synthesis by HF. These upregulatory effects of PLM were abrogated by pretreatment of the cells with a tyrosine kinase inhibitor. These results indicate that PLM upregulates signal transduction in a variety of cell types and the upregulation may induce pulmonary fibrosis.

Increase of Candida cell virulence by anticancer drugs and irradiation.

The influence of anticancer drugs and irradiation on Candida cell proliferation, adherence to HeLa cells and susceptibility to antifungal drugs (amphotericin B and miconazole) and neutrophils were examined using two Candida albicans strains. After treatment with 5-fluorouracil (25 microg/ml to 250 microg/ml), cis-diammine-dichloroplatinum (10 microg/ml to 100 microg/ml), peplomycin (0.5 microg/ml to 5 microg/ml) or 137Cs (20 Gy to 40 Gy) for 3 days or more, surviving Candida cells proliferated more rapidly than did untreated control cells. Anticancer agent-pretreated Candida cells revealed an increased adhesion to HeLa cells corresponding to an increase of binding to the lectins. The concentration of half limited colony formation (IC50) of amphotericin B and miconazole was increased to near two-fold that of the control by pretreatment of Candida cells with the anticancer agents, except peplomycin, which only weakly increased IC50. In addition, the enolase and Candida acid proteinase activities in the culture supernatants were increased by pretreatment with the drugs and irradiation. Correspondingly, surviving Candida cells after these treatments were resistant to neutrophils, with a reduction to half of the killing. These results indicate that anti-cancer drugs and irradiation potentiate the virulence of Candida cells, or they eliminate Candida cells with low virulence, thereby enhancing the risk of oral and systemic candidiasis.

Influence of aging on candidal growth and adhesion regulatory agents in saliva.

Although oral candidiasis is frequently seen in the elderly, the factors determining candidal growth have insufficiently been explored. Hence, we examined the influence of aging on candidal adhesion and growth-inhibitory agents in saliva in 45 healthy volunteers and 60 patients with oral candidiasis. Both non-stimulated and stimulated salivary flow rates (SFRs) in the healthy controls decreased with aging. A gradual decrease of SFRs with aging was also observed in the patients, and the SFR levels were markedly lower than those in the controls. Although the salivary glucose levels were almost constant in all age groups, secretory immunoglobulin A and lactoferrin levels in saliva were significantly decreased statistically with age, and a marginal age-associated decrease in transferrin levels was also observed. In addition, the generation of superoxide from neutrophils in saliva and their Candida killing activity decreased with age, and these phenomena were more apparent in the patients. Furthermore, a larger number of Candida adhered to oral keratinocytes obtained from the elderly healthy controls than to those obtained from young controls. Correspondingly, keratinocytes from the aged controls showed more concanavalin-A binding sites than those from the young controls. However, oral Candida did not increase with increasing age in the controls, although an age-associated increase of oral Candida was observed in the patients. Taken together, these results indicate that the decreases of SFRs and salivary anti-candidal factors, suppression of salivary neutrophil function and the increase of candidal adhesion sites on keratinocytes predispose elderly individuals to oral candidiasis.

Lactoferrin peptide increases the survival of Candida albicans-inoculated mice by upregulating neutrophil and macrophage functions, especially in combination with amphotericin B and granulocyte-macrophage colony-stimulating factor.

To develop a new strategy to control candidiasis, we examined in vivo the anticandidal effects of a synthetic lactoferrin peptide, FKCRRWQWRM (peptide 2) and the peptide that mimics it, FKARRWQWRM (peptide 2'). Although all mice that underwent intraperitoneal injection of 5 x 10(8) Candida cells with or without peptide 2' died within 8 or 7 days, respectively, the survival times of mice treated with 5 to 100 microg of intravenous peptide 2 per day for 5 days after the candidal inoculation were prolonged between 8.4 +/- 2.9 and 22.4 +/- 3.6 days, depending on the dose of peptide 2. The prolongation of survival by peptide 2 was also observed in mice that were infected with 1.0 x 10(9) Candida albicans cells (3.2 +/- 1.3 days in control mice versus 8.2 +/- 2.4 days in the mice injected with 10 microg of peptide 2 per day). In the high-dose inoculation, a combination of peptide 2 (10 microg/day) with amphotericin B (0.1 microg/day) and granulocyte-macrophage colony-stimulating factor (GM-CSF) (0.1 microg/day) brought prolonged survival. With a combination of these agents, 60% of the mice were alive for more than 22 days. Correspondingly, peptide 2 activated phagocytes inducing inducible NO synthase and the expression of p47(phox) and p67(phox), and peptide 2 increased phagocyte Candida-killing activities up to 1.5-fold of the control levels upregulating the generation of superoxide, lactoferrin, and defensin from neutrophils and macrophages. These findings indicated that the anticandidal effects of peptide 2 depend not only on the direct Candida cell growth-inhibitory activity, but also on the phagocytes' upregulatory activity, and that combinations of peptide 2 with GM-CSF and antifungal drugs will help in the development of new strategies for control of candidiasis.

Pulmonary infiltration with eosinophilia (PIE) syndrome induced by antibiotics, PIPC and TFLX during cancer treatment.

Drugs induce a variety of pulmonary diseases including pulmonary infiltration with eosinophilia (PIE) syndrome. We report a case of PIE syndrome which was observed after neck dissection. An 83-year-old male patient attended our clinic complaining of upper neck swelling and was diagnosed as advanced lymph node metastasis related to previously resected oral carcinoma and underwent neck dissection. Despite administration of antibiotics (piperacillin sodium, PIPC; and tosufloxacin tosilate, TFLX), fever and an elevation of the c-reactive protein (CRP) level with neutrophilia appeared, and an infiltration shadow was observed in the right lower pulmonary field. With the suspicion of pneumonia, the antibiotics were exchanged for panipenem/betamipron. However, the pulmonary infiltration spread widely, CRP increased to 12.9 mg/dl and severe eosinophilia (23%) was observed a few days after changing the antibiotics. PIE syndrome was suspected, and the patient underwent steroid mini-pulse therapy consisting of methylprednisolone sodium succinate (500 mg) and prednisolone (60 mg). After steroid therapy, the pulmonary condition largely improved. However, about 2 weeks after the start of steroid administration, a fever and a further elevation of CRP were observed with an increase of beta-D-glucan in serum. Roentgenography revealed diffuse infiltration shadows throughout the lungs, and the patient died about 3 weeks after the onset from respiratory distress. In vitro, blastogenesis of patient's peripheral blood lymphocytes was strongly enhanced by PIPC and TFLX, and they generated a large amount of interleukin-5 in the presence of PIPC or TFLX. The clinical course and laboratory examination results revealed that PIE syndrome may have been induced by PIPC and TFLX and that PIE syndrome should be suspected in treatment of carcinomas when dyspnea and pulmonary infiltration are complicated with eosinophilia.

A novel bovine lactoferrin peptide, FKCRRWQWRM, suppresses Candida cell growth and activates neutrophils.

To identify potent new antifungal agents, the Candida cell growth inhibitory activities of six lactoferrin (Lf) peptides consisting of 6-25 amino acid residues (peptide 1, FKCRRWQWRMKKLGAPSITCVRRAF lactoferricin B; peptide 2, FKCRRWQWRM; peptide 2', FKARRWQWRM; peptide 3, GAPSITCVRRAF; peptide 4, RRWQWR; and peptide 5, RWQWRM) were examined. Of these, peptide 2 strongly suppressed the multiplication of Candida cells, but other peptides showed only weak activities. In two strains of C. albicans, the minimum inhibitory concentration 100 of peptide 2 (17.3+/-2.2 microM and 17.5+/-2.4 microM) was close to that of miconazole (13.0+/-1.7 microM and 13.1+/-1.6 microM) but markedly different from that of amphotericin B (0.52+/-0.09 microM and 0.56+/-0.11 microM). The suppression of Candida cell growth was additively increased by a combination of peptide 2 with amphotericin B and miconazole. Peptides 1, 3, 4 and 5 and Lf suppressed iron uptake by Candida cells, inversely correlated with their Candida cell growth inhibition activities. However, iron uptake was not inhibited by peptide 2. In addition, peptide 2 upregulated Candida cell killing activity of polymorphonuclear leukocytes (PMN) increasing their superoxide generation, protein kinase C activity, p38 MAPK activity and the expression of p47phox. These results indicated that the main antimicrobial activity of the Lf peptides is dependent on the N-terminal half of Lf and that the PMN upregulatory activity of peptide 2 and additive function of peptide 2 with antifungal drugs are useful for prophylaxis and control of candidiasis.

Regulation of cigarette smoke-induced cytochrome P4501A1 gene expression in osteogenic disorder Shionogi rat liver and in lung by large ascorbic acid dose.

The effects of a large ascorbic acid dose on cytochrome P4501A1 gene expression induced by cigarette smoke exposure was studied in Osteogenic Disorder Shionogi rats, which lack ascorbic acid biosynthesis. The rats were divided into four groups and were administered either a minimal amount (4 mg/day, 4S and 4C) or a large amount (40 mg/day, 40S and 40C) of ascorbic acid. The 4S group and 40S group were daily exposed to cigarette smoke for 2 hours, while the 4C group and 40C group were not. At the end of the 25-day experiment, the rats were killed. The cytochrome P4501A1 mRNA level both in the liver and lung was measured by a competitive reverse transcription-polymerase chain reaction method. When a minimal amount of ascorbic acid was administered, the cytochrome P4501A1 mRNA increased in the liver of the cigarette smoke-exposed group (4S) compared with the control group (4C). On the other hand, when a large amount of ascorbic acid was administered, this increase was not observed in the cigarette smoke-induced group (40S) in liver. On the other hand, in lung, an increased mRNA level in 4S group was not decreased by large ascorbic acid administration (40S). This is the first direct mRNA-level evidence of the effects of a large ascorbic acid dose on the gene expression stimulated by cigarette smoke.

Enhanced apoptosis of squamous cell carcinoma cells by interleukin-2-activated cytotoxic lymphocytes combined with radiation and anticancer drugs.

Induction of potent apoptosis is required in cancer therapy. We examined the combination effect of interleukin-2-activated lymphocytes (LAK cells) and anticancer drugs or gamma (gamma)-rays on the induction of apoptosis in an established oral squamous cell carcinoma cell line (OSC-3 cells). By pretreatment of OSC-3 cells with (137)Cs (5 Gy), 5-fluorouracil (5-FU) (0.5 microg/ml) or cis-dichlorodiammine-platinum (CDDP) (5 microg/ml), the activation of bid and caspase-3 by LAK cells was strongly increased and associated with an enhanced degradation of poly-(ADP-ribose) polymerase (PARP) and/or nuclear mitotic apparatus protein (NuMA) and the increased fragmentation of DNA. The LAK cell-enhanced caspase-3 activity in the pretreated OSC-3 cells was decreased to approximately 70% and 40% of the control by the addition of Z-AAD-CMK (a granzyme B inhibitor) and neutralising monoclonal antibody to Fas antigen (alphaFas-IgG), respectively. The combined treatment-induced DNA fragmentation was suppressed by approximately 20% and 30% of the control by the addition of Z-AAD-CMK and alpha Fas-IgG, respectively, in the co-culture system. While Ac-DEVD-CHO (a caspase-3 inhibitor) suppressed the DNA fragmentation levels to approximately half and this was similar to the amount of suppression that was obtained by the addition of both alpha Fas-IgG and Z-AAD-CMK. In addition, LAK cell-activated bid may have increased the intracellular reactive oxygen intermediates (ROI) level and induced a decrease of mitochondrial membrane potential. These influences by LAK cells were enhanced when OSC-3 cells were pretreated with each anticancer drug or (137)Cs. Furthermore, the increase of ROI by LAK cells was suppressed by alpha Fas-IgG and Z-AAD-CMK to approximately half the level of the control. These results indicate that anticancer drugs and gamma-rays prime squamous cell carcinoma cells to be susceptible to apoptosis by LAK cells, that LAK cell-induced apoptosis largely depends on the activation of caspase-3 by the Fas/Fas-ligand signal and granzyme B, and that LAK cells induce ROI in the target cells, which is largely mediated by Fas and granzyme B.

Regulation of Candida albicans growth and adhesion by saliva.

To examine the local regulation of oral Candida albicans growth, we examined non-stimulated and stimulated salivary flow rates (SFRs) and the C. albicans growth and adhesion inhibitory activities of saliva in 60 patients with oral candidiasis (divided into two groups: 25 patients with oral candidiasis only (group OC) and 35 patients with oral candidiasis and systemic diseases (group CS)) and 30 healthy control subjects. Both non-stimulated and stimulated SFRs in patients, especially in group CS; were decreased in comparison with those in the healthy control subjects. The levels of secretory immunoglobulin A (sIgA) in group OC and group CS and the lactoferrin level in group CS were decreased as compared with those in control individuals, although there were no differences in transferrin and total secretory component (SC) levels between the three groups. The secretion amounts (microg/min) of these proteins were statistically significantly decreased in the patients, especially in group CS. Saliva from the patients showed a lesser inhibitory effect on C. albicans growth and adhesion to HeLa cells than did saliva from the control subjects. In addition, polymorphonuclear leukocytes (PMNs) in patients' saliva generated smaller amounts of superoxide than did those in control subjects' saliva, and phagocytic and C. albicans killing activities were suppressed in the patients. These results indicate that the decreases in SFR, secretion of antimicrobial proteins in saliva, and salivary PMN activity are risk factors for oral candidiasis associated with aging and systemic diseases.

Candidiasis may induce glossodynia without objective manifestation.

BACKGROUND: The causes of glossodynia in the absence of objective abnormalities range widely and differential diagnosis of glossodynia is very difficult. METHODS: Based on the examination results of peripheral blood, stimulated and nonstimulated salivary flow rate (SFR), glossal pain threshold, and C. albicans cell culture and the response to treatment, we identified the cause of vague pain of the tongue in 98 patients who lacked objective findings and identified candidiasis as the cause of glossodynia in 26 patients. RESULTS: These patients revealed hyposalivation and decreased glossal pain thresholds and C. albicans cell overgrowth. Pain thresholds in the painful portion (54.6+/-2.9 degrees C) were significantly decreased compared with those in the painless portion (57.7+/-3.4 degrees C) (P < 0.05) and the pain thresholds were largely increased after treatment (57.2+/-1.6 degrees C). Nonstimulated SFR before treatment was lower than that of age- and gender-matched healthy people, although stimulated SFR was decreased only slightly. C. albicans cell overgrowth was detected by the number of C. albicans colonies that formed in Sabouraud's agar plate (539.3+/-198.4/dish). After the subsidence of glossal pain by mouth washing with a 3% amphotericin B solution, the C. albicans colonies were decreased to 31.5+/-19.3/dish, which was almost same as the control level, 14.1+/-8.4/dish. CONCLUSION: These results indicate that candidiasis in conjunction with hyposalivation may induce pain in the tongue without manifestation of objective abnormalities.

The pathophysiology of glossal pain in patients with iron deficiency and anemia.

BACKGROUND: It is well known that prolonged anemia causes atrophy of tongue papillae, glossal pain, and dysphagia, but it is uncertain whether iron (Fe) deficiency induces glossal pain without any objective manifestation. To resolve this matter, the relationship between Fe deficiency and glossal pain was examined. METHODS: Eighteen patients with Fe deficiency and 7 anemic patients manifesting spontaneous irritation or pain of the tongue without any objective abnormalities participated in this study. To ascertain the cause of glossal pain and the oral pathophysiology in Fe deficiency and anemia, peripheral blood was examined and the glossal pain threshold and salivary flow rates (SFRs) were estimated along with Candida albicans cell culture tests. RESULTS: Compared with patients with Fe deficiency, those with anemia had a longer history of tongue pain. In patients with anemia, painful areas of the tongue were more numerous than in patients with Fe deficiency. Pain thresholds were decreased in the painful portions, and both nonstimulated and stimulated SFRs were suppressed. Each patient was treated with oral Fe; within 2 months, most patients exhibited increased serum ferritin level (P< 0.02, paired t-test), pain threshold (P < 0.05) and salivation (P < 0.05) and glossal pain subsided. CONCLUSIONS: Fe deficiency causes glossal pain and the degree of glossal pain increases as Fe deficiency advances to anemia, manifesting hyposalivation and abnormalities of glossal papillae.

Manganese superoxide dismutase negatively regulates the induction of apoptosis by 5-fluorouracil, peplomycin and gamma-rays in squamous cell carcinoma cells.

We investigated the relationship between manganese superoxide dismutase (Mn-SOD) activity and apoptosis induced by anticancer drugs and radiation. Although the activity of copper, zinc-SOD did not differ greatly among 9 squamous cell carcinoma (SCC) cell lines (OSC-1 to OSC-9), the Mn-SOD activity did differ among the cell lines. The Mn-SOD activity was increased by treatments with 5-fluorouracil (5-FU), peplomycin and 137Cs, reaching plateau levels at 12 h after treatment and then decreasing gradually. When OSC-1 and OSC-3, and OSC-2 and OSC-4 were examined as representative cell lines with low and high Mn-SOD activity, respectively, the decrease was more prominent in OSC-1 and OSC-3 than in OSC-2 and OSC-4. The intracellular levels of superoxide and hydrogen peroxide (H2O2) were increased after treatment with the anticancer agents, and the increases were larger in OSC-1 and OSC-3 than in OSC-2 and OSC-4. The decrease of mitochondrial membrane potential (deltapsi(m)) by the anticancer agents was marked in OSC-1 and OSC-3. Correspondingly, the release of cytochrome c, the activation of caspase-3 and the cleavage of poly(ADP-ribose)polymerase were stronger in OSC-3 than in OSC-4. In addition, apoptosis induced by the anticancer agents was prominent in OSC-3, exhibiting a close relationship with the deltapsi(m) and the H2O2 level. These results indicate that Mn-SOD in SCC cells modulates apoptosis induction and the inactivation of Mn-SOD might be a promising strategy for SCC treatment.

The inhibitory action of BOF-A2, a 5-fluorouracil derivative, on squamous cell carcinoma.

Inactivation of antineoplastics is a serious problem in cancer therapy, and prevention of the inactivation is a surpassing strategy for enhancement of their therapeutic effects. BOF-A2, which contains both EM-FU, a masked form of 5-fluorouracil (5-FU), and CNDP, an inhibitor of 5-FU degradation, was developed with this aim. We compared the antitumor effects of BOF-A2 and 5-FU in human squamous cell carcinoma cells transplanted to nu/nu mice. Each drug (0.9-7.0 mg/kg of 5-FU and 3.8-30 mg/kg of BOF-A2) was orally administered every day to 5 mice in each dosage group for 4 weeks. Although the maximal tumor growth inhibition by 5-FU (3.5 mg/kg per day) was about 50% of the control value, 15 mg/kg per day of BOF-A2, which is equimolar to 3.5 mg/kg per day of 5-FU, almost completely inhibited the tumor growth. The flow-cytometric analysis revealed that BOF-A2 (15 mg/kg per day) induced more prominent S-phase-accumulation (63 +/- 6%) of tumor cells than did 3.5 mg/kg per day of 5-FU (43 +/- 18%), and immunohistochemical stainings indicated that the decrease of proliferating cell nuclear antigen expression was more prominent in tumor cells in the BOF-A2-treated mice than in the 5-FU-treated mice. Correspondingly, the DNA synthesis was markedly suppressed in tumor cells obtained from BOF-A2-treated mice. Compared with 5-FU, BOF-A2 more strongly induced apoptosis; apoptotic cells detected by nick-end labeling techniques were about 20% of the tumor cells in 5-FU (3.5 mg/kg per day)-treated mice, and nearly 50% in BOF-A2 (15 mg/kg per day)-administered mice. The expressions of involucrin, cytokeratin 10 and protein kinase C eta, which are associated with squamous cell differentiation, were not increased by BOF-A2 or 5-FU, although the expression of transglutaminase was slightly augmented by both drugs. These results indicate that compared with 5-FU, BOF-A2 more strongly suppresses the growth of squamous cell carcinoma by inhibiting DNA synthesis and inducing apoptosis but not cell differentiation.

A splicing mutation in the hydroxymethylbilane synthase gene in a Japanese family with acute intermittent porphyria.

OBJECTIVES: Acute intermittent porphyria (AIP) is an autosomal dominant inherited disease caused by a decreased activity of hydroxymethylbilane synthase (HMBS). As far as the gene abnormalities of the HMBS, many different mutations have been reported. In this work, we investigated the presence of mutations in a Japanese family with AIP. DESIGN AND METHODS: A 44-year-old Japanese male and nine members of his family were investigated. All of them were screened by traditional biochemical markers. Mutational analysis was performed using polymerase chain reaction-single strand conformation polymorphism method followed by DNA sequencing. A reliable restriction enzyme cleavage assay was established for the pedigree analysis. RESULTS: The mutation was a splicing mutation, a C to G transversion at position -3 of the acceptor site of intron 11 of the HMBS gene, resulting in the exon 12 skipping. The patient is heterozygous for the mutation, and his father appeared to be the source of the mutant allele. This mutation created a new cleavage site of the Nla III restriction enzyme and could be screened by a amplified fragment from genomic DNA with digestion. Using this cleavage assay, an asymptomatic carrier in the family was definitively identified. CONCLUSIONS: This mutation was first found among Japanese AIP patients, but happened to be the same as reported previously from Europe. A similarity of gene abnormality may suggest that those European and Japanese AIP families have a common ancestor. Molecular investigations on the family members should be applied not only for more accurate diagnosis, but also for understanding the molecular genetic heterogeneity underlying this dominantly inherited enzymopathy.