Complex intrinsic abnormalities in osteoblast lineage cells of X-linked hypophosphatemia: Analysis of human iPS cell models generated by CRISPR/Cas9-mediated gene ablation.

X-linked hypophosphatemia (XLH) is caused by inactivating variants of the phosphate regulating endopeptidase homolog X-linked (PHEX) gene. Although the overproduction of fibroblast growth factor 23 (FGF23) is responsible for hypophosphatemia and impaired vitamin D metabolism, the pathogenesis of XLH remains unclear. We herein generated PHEX-knockout (KO) human induced pluripotent stem (iPS) cells by applying CRISPR/Cas9-mediated gene ablation to an iPS clone derived from a healthy male, and analyzed PHEX-KO iPS cells with deletions extending from exons 1 to 3 and frameshifts by inducing them to differentiate into the osteoblast lineage. We confirmed the increased production of FGF23 in osteoblast lineage cells differentiated from PHEX-KO iPS cells. In vitro mineralization was enhanced in osteoblast lineage cells from PHEX-KO iPS cells than in those from isogenic control iPS cells, which reminded us of high bone mineral density and enthesopathy in patients with XLH. The extracellular level of pyrophosphate (PPi), an inhibitor of mineralization, was elevated, and this increase appeared to be partly due to the reduced activity of tissue non-specific alkaline phosphatase (TNSALP). Osteoblast lineage cells derived from PHEX-KO iPS cells also showed the increased expression of multiple molecules such as dentine matrix protein 1, osteopontin, RUNX2, FGF receptor 1 and early growth response 1. This gene dysregulation was similar to that in the osteoblasts/osteocytes of Phex-deficient Hyp mice, suggesting that common pathogenic mechanisms are shared between human XLH and Hyp mice. Moreover, we found that the phosphorylation of CREB was markedly enhanced in osteoblast lineage cells derived from PHEX-KO iPS cells, which appeared to be associated with the up-regulation of the parathyroid hormone related protein gene. PHEX deficiency also affected the response of the ALPL gene encoding TNSALP to extracellular Pi. Collectively, these results indicate that complex intrinsic abnormalities in osteoblasts/osteocytes underlie the pathogenesis of human XLH.

Growth-related skeletal changes and alterations in phosphate metabolism.

Serum inorganic phosphate (Pi) levels are higher in children than in adults; however, the underlying mechanisms remain unclear. Therefore, we herein attempted to elucidate the mechanisms altering Pi metabolism from youth to adulthood using 4-week-old (young) and 12-week-old (adult) mice. Despite higher serum Pi levels, serum fibroblast growth factor 23 (FGF23) levels were lower in young mice, and the amount of FGF23 in bone tended to increase from youth to adulthood. Increases in serum FGF23 levels during growth were associated with the up- and down-regulation of the renal expression of Cyp24a1 encoding vitamin D-24-hydroxylase and Slc34a3 encoding the type IIc sodium/phosphate (Na+/Pi) co-transporter, respectively, suggesting an enhancement in the FGF23-mediated bone-kidney axis from youth to adulthood. We then isolated osteoblasts and osteocytes from young and adult mice and compared the expression of genes involved in Pi metabolism and/or mineralization. In contrast to the growth-related increase in Fgf23 expression, the expression of some genes, including the dentin matrix protein 1 (Dmp1) and phosphate-regulating gene with homologies to endopeptidases on the X chromosome (Phex) markedly decreased from youth to adulthood. The down-regulation of Dmp1 and Phex may contribute to growth-related increases in FGF23. The responses of isolated osteoblasts and osteocytes to high Pi levels also markedly differed between young and adult mice. Treatment of isolated osteocytes with high Pi increased the production of FGF23 in adult mice but not in young mice. These results indicate a close relationship between skeletal changes from youth to adulthood and an alteration in Pi metabolism, and provide insights into the mechanisms by which osteoblasts and osteocytes maintain Pi homeostasis.

Clonal osteoblastic cell lines with CRISPR/Cas9-mediated ablation of Pit1 or Pit2 show enhanced mineralization despite reduced osteogenic gene expression.

Multiple actions of extracellular Pi on the skeletal cells are likely to be partly mediated by type III sodium/phosphate (Na+/Pi) cotransporters Pit1 and Pit2, although the details are not fully understood. In the current study, to determine the roles of Pit1 and Pit2 in osteoblasts, we generated Pit1-knockout (KO) and Pit2-KO osteoblastic cells by applying CRISPR/Cas9 genome editing to an osteoblastic cell line MC3T3-E1 subclone 4. The extracellular Pi level was increased in the Pit1-KO and Pit2-KO clones due to the reduced Pi uptake. Interestingly, in vitro mineralization was accelerated in the Pit1-KO and Pit2-KO clones, although the induction of the expression of osteogenic marker genes was suppressed. In the cells before mineralization, extracellular levels of pyrophosphate (PPi) and adenosine triphosphate (ATP) were increased in the Pit1-KO and Pit2-KO clones, which might be attributable to the reduced expression and activity of tissue-nonspecific alkaline phosphatase (TNSALP). A 24-h treatment with high Pi reduced the expression and activity of TNSALP, suggesting that the suppression of TNSALP in the Pit1-KO and Pit2-KO clones was caused by the increased availability of extracellular Pi. Lentiviral gene transfer of Pit1 and Pit2 restored the changes observed in Pit1-KO and Pit2-KO clones, respectively. The expressions of P2Y2 and P2X7 which encode receptors for extracellular ATP were altered in the Pit1-KO and Pit2-KO clones, suggesting an influence on purinergic signaling. In mineralized cells after long-term culture, intracellular levels of PPi and ATP were higher in the Pit1-KO and Pit2-KO clones. Taken together, ablation of Pit1 or Pit2 in this osteoblastic cell model led to accelerated mineralization, suppressed TNSALP and altered the levels of extracellular and intracellular PPi and ATP, which might be partly mediated by changes in the availability of extracellular Pi.

Development of a brief screening interview for adjustment disorders and major depression in patients with cancer.

BACKGROUND: Adjustment disorders and major depression are common psychiatric disorders in patients with cancer and have a serious impact on quality of life. The problem in clinical oncology settings is underrecognition of these disorders; as a result, screening is recommended to detect them. The goal of the current study was to develop a new, brief screening tool for adjustment disorders and major depression and to compare its performance with that of existing screening methods. METHODS: Patients with cancer completed the newly developed One-Question Interview (a 1-item, structured interview); the Distress Thermometer (a 1-item, self-report questionnaire), which previously was developed as a brief screening tool; and the Hospital Anxiety and Depression Scale (HADS; a 14-item, self-report questionnaire). Psychiatric diagnoses of adjustment disorders and major depression were made by psychiatrists and were based on criteria set forth by the Diagnostic and Statistical Manual of Mental Disorders, 4th edition. RESULTS: Two hundred seventy-five patients participated in the study. Scores on both the One-Question Interview and the Distress Thermometer were significantly correlated with HADS score (One-Question Interview: r = -0.66, P < 0.01; Distress Thermometer: r = 0.71, P < 0.01). At the optimal cutoff points, the sensitivity and specificity for detection of adjustment disorders and major depression were 80% and 61%, respectively, for the One-Question Interview; 84% and 61%, respectively, for the Distress Thermometer; and 92% and 57%, respectively, for the HADS. CONCLUSIONS: The results of the current study suggested that the One-Question Interview was a valid tool for use in screening patients with cancer for adjustment disorders and major depression. Its performance was inferior to that of the HADS but comparable to that of the Distress Thermometer. The One-Question Interview may be suitable for widespread use in routine screening.

Somatic symptoms for diagnosing major depression in cancer patients.

Diagnosing depression in cancer patients has been challenging because the diagnostic criteria include somatic symptoms frequently attributed to the cancer itself or its treatment. However, few studies have explored how to appropriately deal with individual somatic symptoms. The authors used data from 220 cancer patients with major depression to examine the intercorrelations among the DSM-IV somatic and nonsomatic symptom criteria as well as whether the presence of an individual somatic symptom could discriminate the severity of major depression. Appetite changes and a diminished ability to think were positively associated with anhedonia. Sleep disturbance and fatigue were not significantly associated with nonsomatic symptoms. These associations were consistent after adjusting for physical functioning and pain. Only patients with appetite changes showed a higher severity of depression. These results suggest that individual somatic symptoms differ in nature and that appetite-related symptoms and a diminished ability to think may be useful for diagnosing depression in cancer patients, whereas sleep disturbances and fatigue may not be as useful.

Psychiatric evaluation of competency in cancer patients.

This study was conducted to investigate the profile of patients referred for psychiatric evaluation of competency among patients with cancer. Among the 1721 referred cancer patients, 43 (2.5%) were referred for competency evaluation. The most common reason for competency evaluation was refusal of test or treatment, and the common psychiatric diagnoses were adjustment disorders, delirium and personality disorders. Cancer patients with personality disorders were more likely to be evaluated as competent, while patients with delirium and dementia were often incompetent; those with adjustment disorders, major depression and psychotic illness could be either competent or incompetent. While these findings were almost consistent with those reported from previous studies, some results may be unique to cancer patients.

Clinical factors associated with suicidality in cancer patients.

BACKGROUND: Previous epidemiological studies have indicated that the risk of suicide in cancer patients is higher than that of the general population. In addition, euthanasia and physician-assisted suicide (PAS) have recently become controversial medical, ethical and legal issues all over the world. Although suicide in cancer patients and appropriate management of cancer patients with suicidality are critical issues in clinical oncology practice, there have been very few studies to understand suicidality in cancer patients. The purpose of this study was to explore the clinical factors associated with suicidality in Japanese patients with cancer. METHODS: We investigated the clinical factors associated with suicidality in cancer patients by analyzing the consultation data of patients referred to the Psychiatry Division, National Cancer Centre Hospital and Hospital East, Japan. RESULTS: Of 1713 psychiatric referrals, 62 (3.6%) were related to suicidality, including 44 cases with suicidal ideation, 10 suicide attempts and eight cases who had requested euthanasia and/or continuous sedation. Most of the patients suffered from physical distress and/or psychiatric disorders. The results of a multivariate analysis comparing cancer patients with a psychiatric referral related to suicidality and those referred for other reasons indicated that impaired physical functioning and major depression were significant associated factors. CONCLUSIONS: Our findings suggest that early detection and appropriate management of major depression and comprehensive care improving physical functioning may help to prevent suicide and manage suicidality in Japanese cancer patients.

Clinical experience of the pharmacological treatment algorithm for major depression in advanced cancer patients: preliminary study.

INTRODUCTION: Treatment of major depression in advanced cancer patients is often difficult because of their special characteristics. METHOD: The authors developed a treatment algorithm for major depression in advanced cancer patients and report on their clinical experience using it. The applicability, tolerability, and clinical efficacy of the algorithm were evaluated in 95 advanced cancer patients with major depression. RESULTS: The algorithm was not suitable for seven patients and was not used correctly in 14 cases. It was correctly applied to 74 patients (77%), 23 of whom dropped out for cancer-related reasons (deterioration of physical condition, transfer to other hospitals, cancer death). As for tolerability, 22 patients (43%) of the 51 dropped out of the antidepressant treatment regimen because of delirium due to deterioration of their physical condition, adverse effects of the antidepressant, etc. In the 29 cases that could be followed up, clinical efficacy was evaluated for 4 weeks, and improvement was observed in 22 cases (76%). CONCLUSION: These preliminary findings suggest that use of the algorithm may be feasible, but that it requires some alterations to manage major depression in advanced cancer patients. (Int J Psych Clin Pract 2002; 6: 83-89).

Psychological distress of family members with cancer patients in Japan.

BACKGROUND: There were no previous studies in Japan on the psychological distress of members of families with cancer patients which focussed on the disclosure of the diagnosis of cancer. This study was designed to investigate factors that may have an effect on the psychological distress of family members. METHODS: The subjects were 95 members of families of cancer patients in the surgical ward; one member was recruited from each patient's family. The psychiatrist investigated the demographic factors of both the patient and the family member: for the patient - gender, age, occupation, cancer site, disclosure (or not) of cancer diagnosis, cancer stage and performance status (PS); for the family member - gender, age, occupation, relationship to the patient, physical illness, frequency of visiting the ward, the period from when the family member was informed of the diagnosis, and any past experience of the loss of close relatives due to cancer. Furthermore, we conducted a survey on the family member's anxiety and depression by using the Spielberger State - Trait Anxiety Inventory (STAI) and the Center for Epidemiological Studies Depression Scale (CES-D). RESULTS: A multiple regression analysis indicated that the factors which were associated with the STAI scores independently were the lack of disclosure of the diagnosis to the patient (P=0.01), and advanced or recurrent cancer (P=0.01). The factors which were associated with the CES-D scores independently were the lack of disclosure of the diagnosis to the patient (P=0.03), advanced or recurrent cancer (P=0.01), and the family member's past history of psychiatric disorders (P=0.01). CONCLUSIONS: The results suggested that the psychological distress of a family member increases when the patient is not informed of the cancer diagnosis, when the cancer is advanced or recurrent, and when the family member has a past history of psychiatric disorders. (Int J Psych Clin Pract 2002; 6: 205-210 ).