Natsuko Tokushige

University of Sydney, Sydney, New South Wales, Australia

Are you Natsuko Tokushige?

Claim your profile

Publications (26)87.93 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Purpose: To assess endometrial gene as well as protein expression of neuroendocrine and supposedly endometriosis-associated product PGP9.5 and pain symptoms in women with endometriosis and controls undergoing laparoscopy, using molecular biological and immuno-histochemical approaches in the same patients. Methods: Biopsy of eutopic endometrium from 29 patients by sharp curettage, and preparation of paraffin blocks. Determination of PGP9.5 gene expression and protein abundance using qPCR and immuno-histochemistry. Results: qPCR; The PGP9.5 mRNA expression level between women with (N = 16) and without (N = 13) endometriosis was not different, regardless of pain symptoms or menstrual cycle phase. PGP9.5 expression was higher in women who reported pain compared to those who did not; however, this association was not statistically significant. The expression of PGP9.5 mRNA was higher in women with endometriosis and pain during the proliferative than in the secretory phase (P = 0.03). Furthermore, in the first half of the cycle, the abundance of the PGP9.5 transcript was also significantly higher in endometriosis patients compared to those without (P = 0.03). Immuno-histochemistry; Thirteen of the 16 endometriosis patients showed positive PGP9.5 immuno-reactivity in the endometrium, whereas no such signal was observed in women without endometriosis. The absolute number of nerve fibres per mm(2) in women with endometriosis was similar, regardless of the pain symptoms. Conclusions: PGP9.5 mRNA expression is increased in the proliferative phase of endometriotic women with pain. The presence of nerve fibres was demonstrated by a PGP9.5 protein signal in immuno-histochemistry and restricted to patients with endometriosis. Based on these results, however, there did not appear to be a direct association between the gene expression and protein abundance in women with and without endometriosis or those that experienced pain.
    No preview · Article · Jul 2014 · Archives of Gynecology and Obstetrics
  • [Show abstract] [Hide abstract]
    ABSTRACT: Endometriosis has been recognized as the principal cause of pelvic pain in most modern industrialized societies for the past 20 years, but mechanisms are remarkably poorly understood. There is great variability in the types and severity of pelvic pain experienced by women with endometriosis, and this is probably a reflection of a variable genetic background enhanced by highly variable environmental and reproductive experiences. Individual women exhibit great variability in perception and tolerance of pain experiences and in the possible development of hyperalgesia, allodynia and even neuropathic pain. The uterus (and especially the eutopic endometrium) and ectopic endometriotic lesions are richly innervated with a range of sensory and autonomic nerve fibers in a way which is never seen in women without endometriosis. These tissues show abundant expression of nerve growth factor, other neurotropins and their receptors, molecules that usually play a central role in sensitizing nociceptors to potential pain stimuli, as well as stimulating nerve fiber growth. Genetic control of secretion and function of neurotropins and the involvement of immune cells, such as macrophages, dendritic cells, B-cells, natural killer cells, mast cells and regulatory T-cells in controlling neurotropins are fields of current active research. Little is known about these mechanisms, other types of molecular stimuli and the processes influencing perception of chronic pelvic pain in endometriosis. A better understanding of these mechanisms will undoubtedly lead to more specific, and hopefully more effective, approaches to management of these very distressing symptoms.
    No preview · Article · Jan 2012

  • No preview · Article · Jun 2011 · Pathology
  • Guoyun Wang · Natsuko Tokushige · Ian S Fraser
    [Show abstract] [Hide abstract]
    ABSTRACT: There was no difference in the density of nerve fibers across the menstrual cycle in peritoneal endometriotic lesions. These findings may explain why patients with peritoneal endometriosis often have painful symptoms throughout the menstrual cycle.
    No preview · Article · Feb 2011 · Fertility and sterility
  • [Show abstract] [Hide abstract]
    ABSTRACT: Endometriosis is a common gynecological condition, characterized by presence of endometrial-like tissue outside the uterine cavity. It is estimated that endometriosis affects up to 10% of women during their reproductive years, causing debilitating pain and infertility. The etiology of this condition remains poorly understood, however evidence suggests that certain immune factors are almost certainly playing crucial roles in pathogenesis and progression of this inflammatory condition.Numerous studies have shown that the immune response is significantly disturbed in the ectopic endometriotic lesions and the surrounding peritoneal fluid in women with endometriosis. It is likely however, that these immune reactions occur in response to presence of foreign endometrial antigens in the peritoneal cavity and are thus secondary to the disease establishment. In fact, recent findings also suggest that an altered immune response in the eutopic endometrium of women with endometriosis is likely to be a key phenomenon which precedes the well-documented changes in lesions at ectopic sites.To date, the most widely accepted theory of pathogenesis of endometriosis remains the theory of repeated retrograde menstruations. This theory suggests that, while under normal conditions a portion of menstrual reflux escapes through the fallopian tubes into the peritoneal cavity during the process of menstruation, in women with endometriosis these shed endometrial fragments are more viable and have multiple molecular properties which make them more capable of implanting at ectopic sites, establishing the disease. The microanatomy and functions of eutopic endometrium of women with endometriosis are strikingly different from those of women without endometriosis. Components of innate immunity and antigen-specific response appear to be significantly altered in endometriosis. Novel studies have shed new light as how endometrial leukocytes may be unable to effectively target shed endometrial antigens, permitting their survival and establishment of ectopic lesions.This chapter will aim to explore the role of immune response in establishment and progression of endometriosis, with particular focus on the role of antigen presentation and immune regulation. We will explore changes in the immune environment at uterine, peritoneal and systemic levels and better define these responses. Better understanding of the role of immune response in endometriosis will provide insights as to how the immune system tries to fight this disease, why and at which point it fails. Such understanding will ultimately affect how we view this disease, how we diagnose it and with the possible aid of novel immuno-modulation therapies may help with better management in the future.
    No preview · Article · Feb 2011
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: To investigate whether proteins secreted in urine differ between women with and without endometriosis. Laboratory study using human urine. University-based laboratory. Women with and without endometriosis undergoing laparoscopy, hysteroscopy and curettage. Urine collection from women with and without endometriosis. Proteomic techniques and mass spectrometry to identify proteins secreted in the urine of women with and without endometriosis. On average, 133 proteins were significantly different between women with and without endometriosis. Cytokeratin-19 was highly up-regulated in the urine of women with endometriosis. Cytokeratin-19 may be a valuable urinary biomarker for endometriosis.
    Full-text · Article · Jan 2011 · Fertility and sterility
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Background / Purpose: Since no non-invasive diagnosis in diagnosing endometriosis is available, there is often substantial delay in the diagnosis of endometriosis and the length of time from the onset of symptoms to the diagnosis of this disease averages between 8 and 12 years. This delay allows considerable progression of this debilitating disease.There are several up-regulated and down-regulated proteins in the urine of women with endometriosis compared with women without the disease. Main conclusion: There were many proteins which were differently expressed between women with and without endometriosis. Cytoketarin 19 was significantly highly up-regulated in urine of women with endometriosis.
    Full-text · Conference Paper · Oct 2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Ovarian endometriomas (n = 29) were innervated by mainly sympathetic and sensory fibers. These fibers may be involved in the generation of pain symptoms.
    No preview · Article · Feb 2010 · Fertility and sterility
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Endometriosis is an inflammatory condition, associated with highly dysregulated immune response at both uterine and peritoneal levels. Surprisingly, Foxp3+ regulatory T-cells, which control and suppress a range of immune responses, have not previously been investigated in endometriosis. Immunohistochemical analysis of Foxp3+ cells in 127 eutopic endometrial samples and 59 ectopic peritoneal lesions revealed that these immune cell populations are highly disturbed in women suffering from endometriosis. We showed that Foxp3+ cells remained highly up-regulated during the secretory phase of the menstrual cycle, while at this time their expression is significantly down-regulated in women without endometriosis (P < 0.001). Foxp3+ cells were detected in the stroma of 18 of the 59 peritoneal endometriotic lesions, but not in the surrounding or control peritoneal tissue. We propose that in eutopic endometrium in women with endometriosis Foxp3+ cells decrease the ability of newly recruited immune cell populations to effectively recognize and target endometrial antigens shed during menstruation, allowing their survival and ability to implant in ectopic sites. At these ectopic sites, variable expression of Foxp3+ cells within some peritoneal endometriotic lesions is likely to be linked to the characteristics and stage of individual lesion development and be playing key roles in pathogenesis and progression of this unique condition.
    Full-text · Article · Feb 2010 · Human Reproduction
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Endometriosis is a common gynaecological disease, but the pathogenesis of endometriosis and pathophysiological basis for endometriosis-associated painful symptoms are still uncertain. Little is known about neuroendocrine (NE) cells in the uterus. For this study, 38 premenopausal women with histologically diagnosed ovarian endometrioma or peritoneal endometriosis and 24 women without endometriosis were selected. Biopsy samples from eutopic endometrium were used for immunohistochemical staining to detect synaptophysin (SYN) and neuron-specific enolase (NSE) expression in women with and without endometriosis. There were substantially more NE cells of eutopic endometrium stained with SYN and NSE in women with endometriosis than in those without endometriosis (3.8 +/- 1.8 versus 0.5 +/- 0.7/mm2, P < 0.001, and 2.8 +/- 2.1 versus 0.4 +/- 0.6/mm2, respectively, P < 0.001). These cells were scattered in the epithelium of endometrial glands. At all stages of the menstrual cycle, the densities of NE cells stained with SYN and NSE were greater in women with endometriosis than in those without endometriosis (P < 0.05). These results suggest that NE cells in eutopic endometrium probably play some role in the pathogenesis or symptoms of endometriosis.
    Preview · Article · Nov 2009 · Human Reproduction
  • [Show abstract] [Hide abstract]
    ABSTRACT: To investigate the extent and types of innervation of endometriotic lesions in various regions of the bowel. Retrospective nonrandomized immunohistochemical study (Canadian Task Force classification II-3. University-based laboratory. Thirty-six women undergoing laparoscopy or laparotomy because of deep infiltrating endometriosis in various regions of the bowel, including the sigmoid colon, appendix, and rectum. Immunohistochemical staining of endometriotic specimens with antibodies against protein gene product 9.5, neurofilament, nerve growth factor, nerve growth factor receptors tyrosine kinase receptor A and p75, growth-associated protein 43, substance P, neuropeptide Y, and vasoactive intestinal peptide to demonstrate myelinated, unmyelinated, sensory, and autonomic nerve fibers. There were significantly more nerve fibers in intestinal deep infiltrating endometriosis (mean [SD] 172.6 [94.2]/mm(2)) than in other deep infiltrating endometriotic lesions (e.g., cul-de-sac and uterosacral ligament) (67.6 [65.1]/mm(2); p<.01). Intestinal deep infiltrating endometriosis was innervated abundantly by sensory Adelta,sensory C, cholinergic, and adrenergic nerve fibers. Nerve growth factor, tyrosine kinase receptor A, and p75 were strongly expressed in endometriotic lesions, and growth-associated protein-43 was also strongly expressed in the endometriosis-associated nerve fibers. The hyperinnervation in intestinal deep infiltrating endometriosis may help to explain why patients with this type of lesion have more severe pain.
    No preview · Article · Nov 2009 · Journal of Minimally Invasive Gynecology
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Diagnosis of endometriosis currently requires a laparoscopy and this need probably contributes to the considerable average delay in diagnosis. We have reported the presence of nerve fibres in the functional layer of endometrium in women with endometriosis, which could be used as a diagnostic test. Our aim was to assess efficacy of nerve fibre detection in endometrial biopsy for making a diagnosis of endometriosis in a double-blind comparison with expert diagnostic laparoscopy. Endometrial biopsies, with immunohistochemical nerve fibre detection using protein gene product 9.5 as marker, taken from 99 consecutive women presenting with pelvic pain and/or infertility undergoing diagnostic laparoscopy by experienced gynaecologic laparoscopists, were compared with surgical diagnosis. In women with laparoscopic diagnosis of endometriosis (n = 64) the mean nerve fibre density in the functional layer of the endometrial biopsy was 2.7 nerve fibres per mm(2) (+/-3.5 SD). Only one woman with endometriosis had no detectable nerve fibres. Six women had endometrial nerve fibres but no active endometriosis seen at laparoscopy. The specificity and sensitivity were 83 and 98%, respectively, positive predictive value was 91% and negative predictive value was 96%. Nerve fibre density did not differ between different menstrual cycle phases. Women with endometriosis and pain symptoms had significantly higher nerve fibre density in comparison with women with infertility but no pain (2.3 and 0.8 nerve fibre per mm(2), respectively, P = 0.005). Endometrial biopsy, with detection of nerve fibres, provided a reliability of diagnosis of endometriosis which is close to the accuracy of laparoscopic assessment by experienced gynaecological laparoscopists. This study was registered with the Australian Clinical Trials Registry (ACTR) 00082242 (registered: 12/12/2007). The study was approved by the Ethics Review Committee (RPAH Zone) of the Sydney South West Area Health Service (Protocol number X05-0345) and The University of Sydney Human Research Ethics Committee (Ref. No. 10761) and all women gave their informed consent for participation.
    Full-text · Article · Sep 2009 · Human Reproduction
  • Source
    Guoyun Wang · Natsuko Tokushige · Robert Markham · Ian S Fraser
    [Show abstract] [Hide abstract]
    ABSTRACT: Deep infiltrating endometriosis (DIE) is a specific type of endometriosis, which can be associated with more severe pelvic pain than other forms of endometriotic lesions. However, the mechanisms by which pain is generated are not well understood. DIE (n = 31) and peritoneal endometriotic (n = 40) lesions were sectioned and stained immunohistochemically with antibodies against protein gene product 9.5, neurofilament, nerve growth factor (NGF), NGF receptors tyrosine kinase receptor-A (Trk-A) and p75, substance P, calcitonin gene-related peptide, vesicular acetylcholine transporter, neuropeptide Y, vasoactive intestinal peptide and tyrosine hydroxylase to demonstrate myelinated, unmyelinated, sensory and autonomic nerve fibres. There were significantly more nerve fibres in DIE (67.6 +/- 65.1/mm(2)) than in peritoneal endometriotic lesions (16.3 +/- 10.0/mm(2)) (P < 0.01). DIE was innervated abundantly by sensory Adelta, sensory C, cholinergic and adrenergic nerve fibres; NGF, Trk-A and p75 were strongly expressed in endometriotic glands and stroma of DIE. The rich innervation of DIE may help to explain why patients with this type of lesion have severe pelvic pain.
    Full-text · Article · Apr 2009 · Human Reproduction
  • [Show abstract] [Hide abstract]
    ABSTRACT: Endometrial polyps are benign lesions frequently identified in women with infertility or abnormal uterine bleeding in the reproductive and postmenopausal phases We report the striking observation that the numbers of activated mast cells expressing tryptase are increased more than sevenfold throughout the cycle in endometrial polyps (n = 20) compared with normal endometrium. This novel finding has important implications for growth, development, and symptoms associated with polyps in many different tissues.
    No preview · Article · Apr 2009 · Fertility and sterility
  • [Show abstract] [Hide abstract]
    ABSTRACT: Immune alterations may be involved in the pathogenesis and progression of endometriosis. Dendritic cells (DCs) are potent antigen presenting cells that are highly involved in the initiation of the immune response. The aim of this study was to investigate DC populations in the eutopic and ectopic endometrium of women with endometriosis compared with controls. Hysterectomy samples were obtained from premenopausal women with (n = 33) and without (n = 28) endometriosis. In addition, paired peritoneal endometriotic lesions and uterine curettings were collected from 32 women with endometriosis. Specimen sections were stained immunohistochemically using antibodies for monoclonal mouse antibodies directed against human CD1a and CD83, which are specific for immature and mature DCs, respectively. The mean density of endometrial CD1a+ DCs in the basal layer was significantly increased in women with endometriosis compared with controls during the proliferative phase only (P = 0.001). There was a highly significant decrease in the density of endometrial CD83+ DCs in women with endometriosis compared with controls in both layers of the endometrium across all phases of the menstrual cycle (P = 0.001). The density of CD1a+ DCs was significantly increased in peritoneal endometriotic lesions (P = 0.003) and in the surrounding peritoneum (P = 0.001) compared with paired uterine curettings and peritoneum distant from the lesion. Both CD1a+ and CD83+ DC populations were altered in the eutopic and ectopic endometrium of women with endometriosis compared with controls. Alterations in these cells, which play a crucial role in the coordination of the immune response, may be involved in pain generation and the pathogenesis of endometriosis.
    No preview · Article · Apr 2009 · Human Reproduction
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Endometriosis is considered to be an inflammatory disease, and macrophages are the most numerous immune cells in endometriotic lesions. However, the mechanisms underlying the elevation of macrophages and their role in the pathogenesis and manifestations of endometriosis still remain unclear. The number of macrophages stained for CD68 in endometriotic lesions (n = 24) and in peritoneum distant from the lesions (n = 14) from women with endometriosis was compared with the number of macrophages in normal peritoneum from women without endometriosis (n = 18). Peritoneal lesions were also double-stained for CD68 and protein gene product 9.5 to study the relationship between macrophages and nerve fibres. The densities of macrophages in peritoneal endometriotic lesions and unaffected peritoneum from women with endometriosis were both significantly higher than that in normal peritoneum from women without endometriosis (P < 0.001). More nerve fibres were also found in the areas where increased numbers of macrophages were identified. There was a significant elevation of macrophages in both normal peritoneum and peritoneal lesions from women with endometriosis compared with normal peritoneum from women without endometriosis. These cells may well play roles in the growth and development of endometriotic lesions and in the generation of pain through interaction with nerve fibres.
    Full-text · Article · Feb 2009 · Human Reproduction
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Endometriosis is an inflammatory condition, characterized by the presence of endometrial-like tissue outside the uterus. The immune system provides a defence mechanism in response to foreign pathogens, and macrophages play important roles in this response. Activation of macrophages has been reported in peritoneal fluid and ectopic endometriotic lesions; however, controversy exists regarding the composition and function of macrophage populations in eutopic endometrium of women with and without endometriosis. This study aimed to quantify macrophages in eutopic endometrium of women with and without endometriosis, during the early, mid and late proliferative and menstrual phases of the cycle. Paraffin-embedded endometrial curettage blocks were selected from pathology archives. Seventy-six specimens from women with and without endometriosis were analysed using standard immunohistochemical techniques with CD68-PGM1 (phosphoglucomutase 1) clone antibody. Macrophages were counted according to their morphology over several fields of view. A significant increase in macrophage cell numbers was shown in eutopic endometrium in women with endometriosis (mean +/- SD, 182.7 +/- 72.9/mm(2)) during all stages of the proliferative phase compared with normal controls (101.6 +/- 53.4/mm(2); P < 0.001). Significant increase in macrophage density occurred in the control group during the mid-menstrual phase, Days 3-4 (P < 0.01), which was not observed in women with endometriosis. This study further supports an association between immune changes in eutopic endometrium and presence of endometriosis. However, it remains uncertain if eutopic immune changes are primary or secondary occurrences.
    Full-text · Article · Feb 2009 · Human Reproduction
  • [Show abstract] [Hide abstract]
    ABSTRACT: Endometriosis is a disease that still presents many puzzles to clinicians and basic research scientists. Until recently, it has been regarded as a condition that arises when normal endometrium adheres to the peritoneal surface and then grows into an `inflammatory' lesion, which adversely influences local reproductive tract function and causes pain. It is now becoming clear that the endometrium within the uterus in these women differs significantly in function from `normal', and that these anomalies probably precede the development of classical ectopic endometriotic lesions. However, the etiology and mechanisms by which endometriosis arises are still far from certain. This review attempts to address the extensive, but fragmented, evidence that demonstrates widespread molecular disturbances in endometrial function and microstructure underlying this complex condition. The review also addresses some of the novel concepts that are being raised by these exciting new discoveries.
    No preview · Article · Dec 2008 · Expert Review of Obstetrics & Gynecology
  • Natsuko Tokushige · Robert Markham · Peter Russell · Ian S Fraser
    [Show abstract] [Hide abstract]
    ABSTRACT: To investigate how progestogens and combined oral contraceptives change nerve fiber density in peritoneal endometriotic lesions and to identify the types of nerve fibers still present during hormone treatment. Laboratory study using human tissue. University-based laboratory. Hormonally treated and untreated women with endometriosis undergoing laparoscopy, hysteroscopy, and curettage. Biopsy samples from peritoneal endometriotic lesions in hormonally treated and untreated women with endometriosis. Types and density of nerve fibers were immunohistochemically determined in peritoneal endometriotic lesions from hormonally treated and untreated women with endometriosis. The nerve fiber density (mean +/- standard deviation/mm(2)) in peritoneal endometriotic lesions from hormone-treated women with endometriosis (10.6 +/- 2.2/mm(2)) was statistically significantly lower than in peritoneal endometriotic lesions from untreated women with endometriosis (16.3 +/-10.0/mm(2)). Nerve growth factor and nerve growth factor receptor p75 expression in peritoneal endometriotic lesions were slightly reduced in hormone-treated women with endometriosis compared with untreated women with endometriosis. Progestogens and combined oral contraceptives reduced nerve fiber density and nerve growth factor and nerve growth factor receptor p75 expression in peritoneal endometriotic lesions.
    No preview · Article · Nov 2008 · Fertility and sterility
  • Ian Stewart Fraser · Natsuko Tokushige · Robert Markham · Peter Russell

    No preview · Article · Jul 2008 · Fertility and sterility