Patient characteristics, diagnostic testing, and initial treatment profiles of patients with connective tissue disease-associated pulmonary arterial hypertension (CTD-PAH) using a Japanese claims database.

OBJECTIVES: To investigate the diagnostic and therapeutic landscape for patients with connective tissue disease (CTD) and CTD-associated pulmonary arterial hypertension (CTD-PAH) in acute-care general hospitals in Japan. METHODS: We conducted a retrospective cohort study by analysing the Medical Data Vision (MDV) database from April 2008 and September 2020. CTD patients who prescribed immunosuppressants were included in cohort 1, and CTD-PAH patients extracted from cohort 1 were included in cohort 2. Patient characteristics, diagnostic screening frequencies for PAH, and initial PAH-specific treatment patterns were assessed. RESULTS: Overall, 16648 patients with CTD and 81 patients with CTD-PAH were included in cohorts 1 and 2, respectively. The frequencies of screening tests for PAH, including brain natriuretic peptide (BNP), transthoracic echocardiogram (TTE), and diffusing capacity of the lungs for carbon monoxide (DLCO), among CTD patients were 0.7, 0.3, and 0.1 tests/person-year, respectively. The most common initial PAH-specific treatment therapy was monotherapy (87.7%), followed by dual therapy (7.4%), and triple therapy (2.5%). CONCLUSION: This is the first study to describe the patient flow from PAH diagnosis to initial PAH-specific treatment for real-world patients who were followed regularly due to CTD in Japanese clinical practice.

Real-world diagnostic landscape and incidence of pulmonary hypertension in adult congenital heart disease patients using administrative claims data in Japan.

BACKGROUND: Although pulmonary hypertension (PH) and Eisenmenger's syndrome (ES) are common complications in adult congenital heart disease (ACHD), the frequency of diagnostic tests and the incidence of PH/ES in patients with ACHD in Japanese clinical practice are unclear. Therefore, we sought to clarify the frequency of diagnostic tests and incidence of PH/ES in patients with ACHD using the Medical Data Vision (MDV) database, the largest anonymized database of diagnosis procedure combination hospitals in Japan. METHODS: We conducted a retrospective cohort study using the MDV database (April 2008 to December 2021) of patients with ACHD (International Classificaiton of Diseases, 10th revision codes: Q203-204, Q210-213, Q250) aged ≥15 years. The frequency of laboratory/clinical tests and the incidence of PH/ES were calculated. Subgroup analyses were performed for the periods 2008-2015 and 2016-2021. RESULTS: Overall, 28219 ACHD patients were extracted from the MDV database (females 56.3%, males 43.7%; mean ± standard deviation age 44.7 ± 23.5 years). The mean ± standard deviation follow-up period was 2.5 ± 2.7 years. The frequencies of electrocardiography, ultrasonography, brain natriuretic peptide (BNP), N-terminal pro-BNP (NT-proBNP), right heart catheterization, and pulmonary function tests (DLCO) were 2149.8, 1054, 1233, 340, 40.0, and 6.0 per 1000 person-years, respectively. The incidence rate of PH/ES was 32.8 per 1000 person-years. The incidence rate of PH/ES increased from 24.6 to 46.7 per 1000 person-years from 2008-2015 to 2016-2021. CONCLUSION: We have clarified the frequency of diagnostic tests related to PH/ES and the incidence of PH/ES in patients with ACHD in clinical practice in Japan, including non-specialist institutions for PH.

Detection of the pan neuronal marker PGP9.5 by immuno-histochemistry and quantitative PCR in eutopic endometrium from women with and without endometriosis.

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.

Nerve fibers and menstrual cycle in peritoneal endometriosis.

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.

Discovery of a novel biomarker in the urine in women with endometriosis.

OBJECTIVE: To investigate whether proteins secreted in urine differ between women with and without endometriosis. DESIGN: Laboratory study using human urine. SETTING: University-based laboratory. PATIENT(S): Women with and without endometriosis undergoing laparoscopy, hysteroscopy and curettage. INTERVENTION(S): Urine collection from women with and without endometriosis. MAIN OUTCOME MEASURE(S): Proteomic techniques and mass spectrometry to identify proteins secreted in the urine of women with and without endometriosis. RESULT(S): 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. CONCLUSION(S): Cytokeratin-19 may be a valuable urinary biomarker for endometriosis.

Nerve fibers in ovarian endometriomas.

Ovarian endometriomas (n = 29) were innervated by mainly sympathetic and sensory fibers. These fibers may be involved in the generation of pain symptoms.

The role of Foxp3+ regulatory T-cells in endometriosis: a potential controlling mechanism for a complex, chronic immunological condition.

BACKGROUND: 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. METHODS AND RESULTS: 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. CONCLUSIONS: 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.

Neuroendocrine cells in eutopic endometrium of women with endometriosis.

BACKGROUND: 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. METHODS: 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. RESULTS: 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). CONCLUSIONS: These results suggest that NE cells in eutopic endometrium probably play some role in the pathogenesis or symptoms of endometriosis.

Hyperinnervation in intestinal deep infiltrating endometriosis.

STUDY OBJECTIVE: To investigate the extent and types of innervation of endometriotic lesions in various regions of the bowel. DESIGN: Retrospective nonrandomized immunohistochemical study (Canadian Task Force classification II-3. SETTING: University-based laboratory. PATIENTS: 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. INTERVENTIONS: 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. MEASUREMENTS AND MAIN RESULTS: 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. CONCLUSION: The hyperinnervation in intestinal deep infiltrating endometriosis may help to explain why patients with this type of lesion have more severe pain.

Dendritic cell populations in the eutopic and ectopic endometrium of women with endometriosis.

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Novel finding of high density of activated mast cells in endometrial polyps.

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.

Macrophages and nerve fibres in peritoneal endometriosis.

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Rich innervation of deep infiltrating endometriosis.

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: The rich innervation of DIE may help to explain why patients with this type of lesion have severe pelvic pain.

Effect of progestogens and combined oral contraceptives on nerve fibers in peritoneal endometriosis.

OBJECTIVE: 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. DESIGN: Laboratory study using human tissue. SETTING: University-based laboratory. PATIENT(S): Hormonally treated and untreated women with endometriosis undergoing laparoscopy, hysteroscopy, and curettage. INTERVENTION(S): Biopsy samples from peritoneal endometriotic lesions in hormonally treated and untreated women with endometriosis. MAIN OUTCOME MEASURE(S): Types and density of nerve fibers were immunohistochemically determined in peritoneal endometriotic lesions from hormonally treated and untreated women with endometriosis. RESULT(S): 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. CONCLUSION(S): Progestogens and combined oral contraceptives reduced nerve fiber density and nerve growth factor and nerve growth factor receptor p75 expression in peritoneal endometriotic lesions.

Macrophage expression in endometrium of women with and without endometriosis.

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

Effects of hormonal treatment on nerve fibers in endometrium and myometrium in women with endometriosis.

OBJECTIVE: To investigate how hormonal treatment can change nerve fiber density and to identify types of nerve fibers in endometrium and myometrium in women with endometriosis. DESIGN: Laboratory study using human tissue. SETTING: University-based laboratory. PATIENT(S): Hormonally treated and untreated women with endometriosis undergoing hysterectomy or curettage. INTERVENTION(S): Endometrial and myometrial tissues were prepared from women with hormonally treated endometriosis and women with untreated endometriosis. MAIN OUTCOME MEASURE(S): Types and density of nerve fibers in endometrium and myometrium in women with hormonally treated and untreated endometriosis were determined immunohistochemically. RESULT(S): The nerve fiber density (mean density +/- SD per square millimeter) in the functional and the basal layers of endometrium (0.2 +/- 0.7/mm(2) and 0.9 +/- 1.3/mm(2), respectively) and myometrium (1.5 +/- 0.8/mm(2)) from women with hormonally treated endometriosis was much lower than that of endometrium (functional layer: 11 +/- 5/mm(2), basal layer: 18 +/- 8/mm(2), respectively) and myometrium (3 +/- 1/mm(2)) from women with untreated endometriosis. Nerve growth factor and nerve growth factor receptor p75 expression was also significantly reduced in women with hormonally treated endometriosis compared with women with untreated endometriosis. CONCLUSION(S): Hormonal treatment significantly reduced nerve fiber density in endometrium and myometrium in women with endometriosis.

A pilot study to evaluate the relative efficacy of endometrial biopsy and full curettage in making a diagnosis of endometriosis by the detection of endometrial nerve fibers.

OBJECTIVE: The purpose of this study was to evaluate endometrial biopsy and curettage in detecting small nerve fibers in eutopic endometrium for diagnosis of endometriosis. STUDY DESIGN: Endometrial biopsies with precise, consistent technique and curettings were taken from 37 women (20 with endometriosis and 17 without endometriosis). Sensitivity, specificity, and positive and negative predictive value were formally calculated. Endometrial nerve fibers were immunohistochemically detected using the pan-neuronal marker PGP9.5. RESULTS: Small nerve fibers were detected in all endometrial biopsies and curettings from all 20 women with endometriosis, but were not detected in endometrium taken from 17 women without endometriosis. Mean (+/-SD) nerve fiber density in the endometrial biopsies was 26.8 per mm(2) +/- 55.9 (range, 1.6-125) and for curettings was 21.6 per mm(2) +/- 33.1 (range, 0.8-250), with 100% specificity, sensitivity, and positive and negative predictive value. CONCLUSION: Careful endometrial biopsy combined with immunohistochemical staining for nerve fibers may be a reliable means of diagnosing or excluding endometriosis.

Different types of small nerve fibers in eutopic endometrium and myometrium in women with endometriosis.

OBJECTIVE: To investigate types of nerve fibers in endometrium and myometrium in women with endometriosis. DESIGN: Laboratory study using human tissue. SETTING: University-based laboratory. PATIENT(S): Women with and without endometriosis undergoing hysterectomy. INTERVENTION(S): Histologic sections of contiguous endometrial and myometrial tissues were prepared from hysterectomies performed on women with and without endometriosis. MAIN OUTCOME MEASURE(S): Types and density of nerve fibers in endometrium and myometrium in women with and without endometriosis were determined using a series of specific markers for neuronal structure and function: PGP9.5, NF, SP, CGRP, TH, VAChT, VIP, and NPY. RESULT(S): Nerve fibers stained with PGP9.5 and NF in endometrium and myometrium were significantly increased in women with endometriosis compared with women without endometriosis. Nerve fibers in the functional layer of endometrium in women with endometriosis were likely to be sensory C, a mixture of sensory A delta, sensory C, and adrenergic fibers in the basal layer of the endometrium, a mixture of sensory A delta, sensory C, adrenergic and cholinergic fibers in the myometrium. CONCLUSION(S): Increased nerve fiber density in endometrium and myometrium, and sensory C fibers and adrenergic nerve fibers in the endometrium in women with endometriosis may play an important role in the mechanisms of pain generation in this condition.