Combined administration of oseltamivir and hochu-ekki-to (TJ-41) dramatically decreases the viral load in lungs of senescence-accelerated mice during influenza virus infection.

To enhance the effect of anti-influenza-virus agent treatment, the effect of combined administration of oseltamivir phosphate and hochu-ekki-to (Japanese traditional herbal medicine, HET) on early viral clearance was examined. Senescence-accelerated mice were given HET in drinking water for 2 weeks, followed by intranasal infection with influenza A virus strain PR8. After 4 hours of infection, oseltamivir was administered orally for 5 days. The viral loads in the lungs of the group receiving combined treatment were dramatically lower when compared with the viral loads in the lungs of the group receiving oseltamivir alone. HET significantly increased the induction of IL-1ß and TNF-α in the lungs of PR8-infected mice and stimulated alveolar macrophage phagocytosis. From these results, we conclude that these functions may be responsible the increased effect on viral load reduction. Here, we show that the combined administration of oseltamivir and HET is very useful for influenza treatment in senescence-accelerated mice.

C/EBPß is involved in the amplification of early granulocyte precursors during candidemia-induced "emergency" granulopoiesis.

Granulopoiesis is tightly regulated to meet host demands during both "steady-state" and "emergency" situations, such as infections. The transcription factor CCAAT/enhancer binding protein ß (C/EBPß) plays critical roles in emergency granulopoiesis, but the precise developmental stages in which C/EBPß is required are unknown. In this study, a novel flow cytometric method was developed that successfully dissected mouse bone marrow cells undergoing granulopoiesis into five distinct subpopulations (#1-5) according to their levels of c-Kit and Ly-6G expression. After the induction of candidemia, rapid mobilization of mature granulocytes and an increase in early granulocyte precursors accompanied by cell cycle acceleration was followed by a gradual increase in granulocytes originating from the immature populations. Upon infection, C/EBPß was upregulated at the protein level in all the granulopoietic subpopulations. The rapid increase in immature subpopulations #1 and #2 observed in C/EBPß knockout mice at 1 d postinfection was attenuated. Candidemia-induced cell cycle acceleration and proliferation of hematopoietic stem/progenitors were also impaired. Taken together, these data suggest that C/EBPß is involved in the efficient amplification of early granulocyte precursors during candidemia-induced emergency granulopoiesis.

Changes in Attitudes of Japanese Doctors toward Complementary and Alternative Medicine-Comparison of Surveys in 1999 and 2005 in Kyoto.

We surveyed the attitudes of Japanese medical doctors toward complementary and alternative medicine (CAM) in 1999. It is supposed that the situation concerning CAM has been changing recently. The aim of the present study is to survey the attitude of doctors toward CAM again, and to examine changes in attitude over the last 6 years. The attitudes of medical doctors belonging to the Kyoto Medical Association toward CAM were surveyed by a structured, self-administered questionnaire in 1999 and 2005. The results showed that the doctors familiar with the term "CAM", practicing CAM therapies, and attending meetings or training courses related with CAM, increased significantly from 1999 to 2005. The doctors who possessed knowledge of CAM also increased significantly from 1999 to 2005. Almost all doctors believed in the effectiveness of Kampo (Japanese traditional herbal medicine) and acupuncture. The number of doctors who believed in the effectiveness of aromatherapy and ayurveda increased significantly in 2005, compared with 1999. In the near future, 58% of doctors desired to practice CAM therapies. In conclusion, the numbers of doctors who practice CAM therapies, possess CAM knowledge and desire to practice such therapies have increased over the last 6 years in Japan.

Isolation of adipose tissue-derived stem cells by direct membrane migration and expansion for clinical application.

Mesenchymal stem cells (MSCs) have recently made significant progression in multiple clinical trials targeting several clinical disorders and in the modulation of immune responses. In the present study, we isolated human adipose tissue-derived stem cells (ADSCs) by direct membrane migration method without using enzymatic digestion via collagenase, and tried to extract adequate number of cells for clinical application. Hydroxyapatite-treated nonwoven fabric membrane made up of synthetic macromolecular fiber materials, polyethylene and polyester terephthalene was used. Expansion culture of ADSCs having plastic flask adherent characteristic in serum-free condition was successfully established, and adequate number of cells were obtained for clinical application. They were found to be positive for CD44, CD73, CD90 and CD105 and negative for CD11b, CD34, CD45, CD80 and HLA-DR. The resulting immunological marker profile satisfied the immunophenotype of previously reported MSCs. Also, microscopic findings demonstrated trilineage differentiation into adipogenic, osteogenic and chondrogenic cells as the characteristics of MSCs. The isolation by nonwoven fabric membrane and expanded cells under serum-free condition satisfied the criteria of MSCs, as proposed by the International Society for Cellular Therapy. Our direct membrane migration method without enzyme digestion is useful as ADSCs can be obtained from small pieces of adipose tissue and expanded under serum-free culture condition. This method was considered to be feasible for clinical application.

MDR1a/1b gene silencing enhances drug sensitivity in rat fibroblast-like synoviocytes.

BACKGROUND: Drug resistance mediated by P-glycoprotein (P-gp) is one of the major reasons for the failure of rheumatoid arthritis (RA) therapy with disease modifying anti-rheumatic drugs and glucocorticoids. In the present study, we aimed to investigate the in vitro effectiveness of small interfering RNA (siRNA) to render rat fibroblast-like synoviocytes (FLS) susceptible to drugs. We also attempted the electroporation-mediated transfer of siRNA against multidrug resistance (MDR) genes into rat knee joints. METHODS: FLS were transfected with siRNAs corresponding to MDR1a and MDR1b genes. FLS were treated with dexamethasone (DEX) and lipopolysaccharide. The mRNA and protein levels of tumor necrosis factor-alpha, interleukin (IL)-6 and IL-1beta were measured. Both siRNAs were co-transduced into rat knee joints by an electroporation method and evaluated the target gene expressions in the synovium. RESULTS: Each siRNA could sequence-specifically reduce the target gene expression by over 70% and effectively suppressed P-gp expression and function in the FLS. Both gene expression and protein production of the inflammatory cytokines in the cells transfected with siRNA were reduced by a greater amount compared to in control cells. The in vivo electroporation-mediated transduction of siRNA could significantly inhibit the target gene expressions. CONCLUSIONS: MDR1a/1b gene silencing by siRNA could effectively inhibit P-gp in rat FLS, resulting in a significant enhancement of the anti-inflammatory effects of DEX. The in vivo siRNA transduction could successfully silence MDR gene expression in the rat synovium. These findings indicate that the siRNA targeting MDR gene could be a useful tool for treating refractory arthritis in RA.

Use of bicistronic vectors in combination with flow cytometry to screen for effective small interfering RNA target sequences.

The efficacy and specificity of small interfering RNAs (siRNAs) are largely dependent on the siRNA sequence. Since only empirical strategies are currently available for predicting these parameters, simple and accurate methods for evaluating siRNAs are needed. To simplify such experiments, target genes are often tagged with reporters for easier readout. Here, we used a bicistronic vector expressing a target gene and green fluorescent protein (GFP) to create a system in which the effect of an siRNA sequence was reflected in the GFP expression level. Cells were transduced with the bicistronic vector, expression vectors for siRNA and red fluorescent protein (RFP). Flow cytometric analysis of the transduced cells revealed that siRNAs for the target gene silenced GFP from the bicistronic vector, but did not silence GFP transcribed without the target gene sequence. In addition, the mean fluorescence intensities of GFP on RFP-expressing cells correlated well with the target gene mRNA and protein levels. These results suggest that this flow cytometry-based method enables us to quantitatively evaluate the efficacy and specificity of siRNAs. Because of its simplicity and effectiveness, this method will facilitate the screening of effective siRNA target sequences, even in high-throughput applications.

Bone marrow angiotensin AT1 receptor regulates differentiation of monocyte lineage progenitors from hematopoietic stem cells.

BACKGROUND: The angiotensin II (Ang II) type 1 (AT(1)) receptor is expressed in bone marrow (BM) cells, whereas it remains poorly defined how Ang II regulates differentiation/proliferation of monocyte-lineage cells to exert proatherogenic actions. METHODS AND RESULTS: We generated BM chimeric apoE(-/-) mice repopulated with AT(1)-deficient (Agtr1(-/-)) or wild-type (Agtr1(+/+)) BM cells. The atherosclerotic development was significantly reduced in apoE(-/-)/BM-Agtr1(-/-) mice compared with apoE(-/-)/BM-Agtr1(+/+) mice, accompanied by decreased numbers of BM granulocyte/macrophage progenitors (GMP:c-Kit(+)Sca-1(-)Lin(-)CD34(+)CD16/32(+)) and peripheral blood monocytes. Macrophage-colony-stimulating factor (M-CSF)-induced differentiation from hematopoietic stem cells (HSCs:c-Kit(+)Sca-1(+)Lin(-)) to promonocytes (CD11b(high)Ly-6G(low)) was markedly reduced in HSCs from Agtr1(-/-) mice. The expression of M-CSF receptor c-Fms was decreased in HSCs/promonocytes from Agtr1(-/-) mice, accompanied by a marked inhibition in M-CSF-induced phosphorylation of PKC-delta and JAK2. c-Fms expression in HSCs/promonocytes was mainly regulated by TNF-alpha derived from BM CD45(-)CD34(-) stromal cells, and Ang II specifically regulated the TNF-alpha synthesis and release from BM stromal cells. CONCLUSIONS: Ang II regulates the expression of c-Fms in HSCs and monocyte-lineage cells through BM stromal cell-derived TNF-alpha to promote M-CSF-induced differentiation/proliferation of monocyte-lineage cells and contributes to the proatherogenic action.

Prevalence of Helicobacter pylori infection in long-term hemodialysis patients.

Patients on hemodialysis often have gastrointestinal complications; however, it is unclear if Helicobacter pylori infection is present in these patients. Here we determined the prevalence of H. pylori infection in 539 Japanese hemodialysis patients by measuring serum anti-H. pylori IgG antibodies. Endoscopy was performed on 299 of these patients and the results were compared to 400 patients with normal renal function who had also undergone endoscopy and sero-testing. A second cohort of 478 dialysis patients, within the original group, was checked serologically for H. pylori infection three times over a four-year observation period. The prevalence of infection in these patients was significantly lower than in those patients with normal renal function, irrespective of the clinical outcomes. The prevalence of H. pylori infection significantly decreased as the duration of dialysis increased, particularly within the first four years following initiation of dialysis. About one-third of patients on dialysis for less than four years became serologically negative for H. pylori infection within this observation period. Our study suggests that although long-term dialysis patients have low prevalence of H. pylori, they still have significant gastroduodenal diseases, such as peptic ulcers, that require endoscopic follow-up.

Asporin and transforming growth factor-beta gene expression in osteoblasts from subchondral bone and osteophytes in osteoarthritis.

BACKGROUND: To clarify the significance of subchondral bone and osteophytes in the pathology of osteoarthritis (OA), we investigated the expression of asporin (ASPN), transforming growth factor-beta1 (TGF-beta1), TGF-beta2, TGF-beta3, and runt-related transcription factor-2 (Runx2) genes involved in bone metabolism. METHODS: Osteoblasts were isolated from 19 patients diagnosed with knee OA and from 4 patients diagnosed with femoral neck fracture. Osteoblast expression of mRNA encoding ASPN, TGF-beta1, TGF-beta2, TGF-beta3, and Runx2 was analyzed using real-time RT-PCR. RESULTS: Expression of ASPN, TGF-beta1, and TGF-beta3 mRNA in the subchondral bone and osteophytes of OA patients increased compared with that of non-OA patients. The ratio of ASPN to TGF-beta1 mRNA in patients with severe cartilage damage was higher than that in patients with mild cartilage damage. CONCLUSIONS: The increased ratio of ASPN mRNA to TGF-beta1 mRNA in patients with severe relative to mild cartilage damage indicates that increased ASPN mRNA expression was significantly associated with the severity of cartilage degeneration. This finding suggests that ASPN may regulate TGF-beta1-mediated factors in the development of OA, which may provide clues as to the underlying pathology of OA.

Influence of extracellular matrix on the expression of inflammatory cytokines, proteases, and apoptosis-related genes induced by hydrostatic pressure in three-dimensionally cultured chondrocytes.

BACKGROUND: The purpose of this study was to investigate the influence of hydrostatic pressure (HP) on the gene expression of cartilage matrix, cytokines, and apoptosis-associated factors in chondrocytes in which the cartilage was in extracellular matrix (ECM)-rich or ECM-poor condition. METHODS: Chondrocytes were isolated from rabbit joints and cultured in alginate beads. Immediately after embedding (0W group) or after 2 weeks culture (2W group), the amounts of glycosaminoglycan (GAG) in the alginate beads were quantified. Both groups were exposed to continuous HP of 10 or 50 MPa for 12 h. The expression of inflammatory cytokines, proteases, and apoptosis-related factors were examined by reverse transcription-polymerase chain reaction (RT-PCR). The expression of proteoglycan core protein (PG) and collagen type II were quantified by real-time RT-PCR. RESULTS: All of the GAG components in alginate beads markedly increased in the 2W group. The expression of PG and collagen type II increased after exposure to 10 MPa in both groups. In the 0W group, these levels decreased after exposure to 50 MPa of HP. The expression of interleukins IL-6 and IL-8 increased after exposure to HP in the 0W group. HP at 50 MPa induced mRNA expression of ADAMTS-5 in the 0W group but not in the 2W group. The expression of Fas increased after exposure to HP in the 0W group. CONCLUSIONS: These findings suggested that nonphysiological, excessive HP on chondrocytes with the ECM in poor condition reduced matrix gene expression and increased expression of the genes associated with apoptosis and catabolism of the cartilage matrix. These results might therefore be associated with the pathogenesis of osteoarthritis.

Nanogel DDS enables sustained release of IL-12 for tumor immunotherapy.

For a valid cytokine immunotherapy of malignancies, a suitable delivery system that ensures slow-release of cytokines is required, because short half-life in vivo of the molecules ruins therapeutic efficacy while causing severe systemic toxic effects. We previously showed that the cholesterol-bearing pullulan (CHP)-based hydrogel nanoparticles, or nanogel, encapsulates, stabilizes and releases various molecules. Here we applied this nanogel to administration in vivo of interleukin-12 (IL-12). Recombinant murine IL-12 (rmIL-12) was successfully incorporated into CHP nanogel simply by incubated with CHP at room temperature. After subcutaneously injected into mice, the CHP/rmIL-12 complex led to a prolonged elevation in IL-12 concentration in the sera. Repetitive administrations of the CHP/rmIL-12, but not rmIL-12 alone, induced drastic growth retardation of preestablished subcutaneous fibrosarcoma without causing any serious toxic event. The present study proposes a novel therapeutic intervention technology, taking advantage of slow and sustained release of bioactive cytokines from the self-assembling biocompatible nanoparticles.

Involvement of IL-17A in the pathogenesis of DSS-induced colitis in mice.

To investigate the etiological implication of IL-17A in inflammatory bowel disease (IBD), dextran sodium sulfate (DSS) was administered to the mice deficient for the IL-17A gene. They showed only faint manifestations of colitis, as revealed by body weight loss, shrinkage in the colon length, serum haptoglobin concentration, and disease activity index. Although the mortality rate of WT mice reached approximately 60%, more than 90% of the IL-17A KO mice survived the DSS treatment. Histological change was also marginal in the IL-17A KO intestine, in which epithelial damage and inflammatory infiltrates were not obvious and the myeloperoxidase activity elevated only slightly. G-CSF and MCP-1 were abundantly produced in WT mouse intestine, whereas the production of these chemokines was drastically hampered in IL-17A-null intestine. The present results show that IL-17A plays a pivotal role in the pathogenesis of DSS-induced colitis, while MCP-1 and G-CSF may be crucially involved in the IL-17A-induced inflammation.

Pleiotrophic functions of Epstein-Barr virus nuclear antigen-1 (EBNA-1) and oriP differentially contribute to the efficiency of transfection/expression of exogenous gene in mammalian cells.

The EBNA1 gene and oriP sequence, originally derived from the EBV genome, provide plasmid vectors with artificial chromosome (AC)-like characteristics, including cytoplasm-to-nuclear transport, nuclear retention, replication and segregation of the DNA, while transcriptional up-regulation has been suggested as another activity of the EBNA1/oriP. Transfection as well as expression rates of various nonviral delivery vehicles are highly improved by inserting these genetic elements into plasmid DNA constructs. Here we differentially analyzed the contribution of each function of the EBNA1/oriP to the efficacy of electroporation-mediated genetic delivery and expression in mammalian cells. It was found that the EBNA1/oriP-mediated acceleration of genetic delivery and expression was predominantly due to the promotion of cytoplasm-to-nuclear recruitment as well as enhancement of transcription, while the episomal replication of the EBV-AC was not essentially involved.

IL-17 is involved in Helicobacter pylori-induced gastric inflammatory responses in a mouse model.

BACKGROUND: Helicobacter pylori (H. pylori) is the major cause of chronic active gastritis and peptic ulcer disease. Recent studies have shown that H. pylori produces various cytokines that are related to neutrophil or mononuclear cell accumulation. Interleukin-17 (IL-17) is the founding member of an emerging family of inflammatory cytokines whose biological activities remain incompletely defined. In this study, the contributions of IL-17 to the induction of gastric inflammation and to the protection from H. pylori infection were investigated using IL-17 gene-knockout (IL-17(-/-)) mice. MATERIALS AND METHODS: IL-17(-/-)and wild-type C57BL/6 mice were challenged with H. pylori CPY2052 (2 x 10(8) CFU/mL) and the histological and microbiological evaluation were carried out at specified times. IL-17 and myeloperoxidase (MPO) protein levels in tissues were assayed in duplicate using ELISA kits. RESULTS: In wild-type mice, IL-17 was undetected at baseline; however, the protein expression of IL-17 was induced after infection with H. pylori. A severe infiltration of neutrophils appeared in the submucosa and the lamina propria in wild-type mice. In contrast, the degree of neutrophil infiltration in IL-17(-/-) mice was significantly lower than that in wild-type mice. Although wild-type mice infected with H. pylori showed drastically higher MPO activity compared with uninfected wild-type mice, any significant increase in the enzyme activity was not revealed in infected IL-17(-/-) mice. The number of H. pylori colonized in the stomach of IL-17(-/-) mice was significantly lower than that of wild-type mice from 1 to 6 months after infection. CONCLUSIONS: These results suggest that IL-17 may play an important role in the inflammatory response to the H. pylori infection and ultimately influence the outcome of the H. pylori-associated disease.

Soluble TRAIL gene and actinomycin D synergistically suppressed multiple metastasis of TRAIL-resistant colon cancer in the liver.

Metastatic liver tumors are highly malignant and refractory to conventional therapies. TRAIL-resistant CT-26 cells underwent apoptosis in vitro in the presence of both recombinant TRAIL (rTRAIL) and a suboptimal dose of actinomycin D (ACD). Co-administration of soluble TRAIL (sTRAIL) gene and ACD suppressed the metastatic liver tumors of CT-26, significantly inducing apoptosis in the tumors, while such effects were not demonstrated in mice that received either the sTRAIL gene or ACD alone. The gene therapy of sTRAIL with a suboptimal dose of an anticancer drug is a new strategy for treatment of multiple liver metastasis.

Sonoporation mediated transduction of pDNA/siRNA into joint synovium in vivo.

The purpose of this study was to investigate the usefulness of sonoporation method on in vivo transduction of plasmid DNA (pDNA) and small interfering RNA (siRNA) into joint tissue. pGEG.GL3 plasmid was mixed with microbubble and injected into knee joints of rats. Ultrasound sonication was performed percutaneously. Three days after injection, GL3 expression of synovial tissue was determined by luciferase assay and RT-PCR. siRNA specific for GL3 (siGL3) or nonspecific siRNA were mixed with pGEG.GL3 plasmid and transduced by sonoporation. siRNA specific for EGFP (siEGFP) was transduced into the knee joints of EGFP transgenic rats, and gene silencing effects for endogenous gene were examined. To determine the localization of transduced siRNA, fluorescently labeled siRNA was transduced into joints. The expression of GL3 in the synovium was significantly enhanced by sonoporation. The gene expression was only seen in the synovium of the knee joint. The expression of GL3 was remarkably suppressed by co-transduction of siGL3, but not suppressed by nonspecific siRNA. siEGFP transduced by sonoporation attenuated green fluorescence on the surface layer of synovium of EGFP transgenic rats. The fluorescently labeled siRNA was seen in the synovium around the patella, femur, and tibia. Sonoporation is examined as a recent, novel, gene transduction method, and the advantage of this technique is minimal invasiveness. In this study, we showed that pDNA/siRNA can be transduced specifically into the joint synovium using sonoporation. The present method may be useful in nucleic acid therapy for joint disorders.

Nationwide survey on complementary and alternative medicine in cancer patients in Japan.

PURPOSE: To determine the prevalence of use of complementary and alternative medicine (CAM) by patients with cancer in Japan, and to compare the characteristics of CAM users and CAM nonusers. PATIENTS AND METHODS: A questionnaire on cancer CAM and the Hospital Anxiety and Depression Scale were delivered to 6,607 patients who were treated in 16 cancer centers and 40 palliative care units. RESULTS: There were 3,461 available replies for a response rate of 52.4%. The prevalence of CAM use was 44.6% (1,382 of 3,100) in cancer patients and 25.5% (92 of 361) in noncancer patients with benign tumors. Multiple logistic regression analysis determined that history of chemotherapy, institute (palliative care units), higher education, an altered outlook on life after cancer diagnosis, primary cancer site, and younger age were strongly associated with CAM use in cancer patients. Most of the CAM users with cancer (96.2%) used products such as mushrooms, herbs, and shark cartilage. The motivation for most CAM use was recommendation from family members or friends (77.7%) rather than personal choice (23.3%). Positive effects were experienced by 24.3% of CAM users with cancer, although all of them received conventional cancer therapy concurrently. Adverse reactions were reported by 5.3% of cancer patients. CAM products were used without sufficient information by 57.3% of users with cancer and without a consultation with a doctor by 60.7% of users. CONCLUSION: This survey revealed a high prevalence of CAM use among cancer patients, without sufficient information or consultation with their physicians. Oncologists should not ignore the CAM products used by their patients because of a lack of proven efficacy and safety.

Combination vaccine of dendritic cells (DCs) and T cells effectively suppressed preestablished malignant melanoma in mice.

The study aims at establishing a novel vaccine procedure, using bone marrow-derived DCs that have ingested apoptotic B16 melanoma (DCs(+)), alone or in combination with splenic T lymphocytes from a syngenic donor. Co-immunization with DCs(+) and T cells showed the highest antitumor potential against preestablished B16 tumor in mice, in which CTL and NK cytotoxicities were drastically elevated, while either DCs(+) alone, naive DCs (DCs(-)) alone, or a mixture of DCs(-) and T cells induced less significant therapeutic outcomes. Use of extracellular matrix proteins elevated antitumor activity of DC(-)/T cell vaccine. Compared with the CD8(+) cells, the CD4(+)T cells more remarkably improved the efficacy of DC-based immunotherapy. The present system may be a feasible therapeutic modality to eradicate malignancies including melanoma.

Hydrostatic pressure induces apoptosis of chondrocytes cultured in alginate beads.

The purpose of this study was to investigate the influence of hydrostatic pressure (HP) on apoptosis and expression of heat-shock protein 70 (HSP70) in chondrocytes cultured in alginate beads. Chondrocytes were isolated from the articular cartilage of rabbit joints and seeded in alginate beads. The beads in Group A were cultured for less than 24 h after being embedded with the chondrocytes, while those in Group B were cultured for 2 weeks. Both groups were exposed to HP of 10 or 50 MPa for 12 or 24 h. The beads in Groups A and B that were not exposed to HP were regarded as controls. Apoptotic cells induced by exposure to HP were quantified using the TUNEL method. Immunohistochemical analysis for HSP70 and in situ TUNEL analysis were also performed. Apoptotic chondrocytes were not observed in the control cells under atmospheric pressure, whereas apoptosis was observed in the beads in Group A, and the number of apoptotic cells increased as the duration and magnitude of HP increased. On the other hand, we observed no significant population of apoptotic cells in the beads in Group B. Chondrocytes expressing HSP70 were not TUNEL positive in the histological analysis. Excessively strong HP could evoke apoptosis when the extracellular matrix did not accumulate around the chondrocytes. HSP70 expression was related to occurrence of apoptosis that resulted from HP. These findings suggest a mechanism for the pathogenesis of cartilage degeneration in osteoarthritis.

Regional Cerebral Blood Flow in [123]I-IMP Single-photon Emission Computed Tomography and the Wechsler Memory Scale-revised in Nondemented Elderly Subjects with Subjective Cognitive Impairment.

Objective Regional cerebral blood flow (rCBF) imaging with single-photon emission computed tomography (SPECT) is useful in the early diagnosis of dementia. We aimed to investigate the association between the rCBF and various domains related to the memory function in elderly subjects with subjective cognitive impairment (SCI). Methods Thirty-two subjects with SCI were included in the present study. Patients with dementia and mild cognitive impairment (MCI) were excluded based on the presence of logical memory impairment. N-isopropyl-p-[123I]-iodoamphetamine SPECT was performed and Wechsler Memory Scale-Revised (WMS-R) was administered to all subjects (mean age, 68.4 years; average Mini-Mental State Examination score, 27.6). The SPECT results were analyzed using the easy Z-score imaging system and the voxel-based stereotactic extraction estimation method. Correlation analyses were performed to investigate the correlation between the mean positive Z-scores in the decrease of the rCBF and the WMS-R indices. Results The SPECT study indicated marked hypoperfusion in some areas, including the bilateral temporal areas, the caudate, and the thalamus, in these subjects in comparison to the normal database. The decrease in the rCBF that was observed in several regions, including the left precuneus and left inferior frontal gyrus (LIFG), showed a significant negative correlation with several indices of the memory function, particularly visual memory. Conclusion The regional hypoperfusion observed in the study using the voxel-based stereotactic extraction estimation method suggest that the regional cerebral dysfunction is associated with the memory function of patients with SCI, even though the subjects in the present study were cognitively intact. The correlation analysis with the WMS-R suggested the contribution of the LIFG to the memory function and indicated the significance of visual memory dysfunction in the neuropsychological assessment to determine the stage of SCI.