Femoral Neck Fractures

Table 1 lists the 35 administrative data case definitions that were selected for investigation. These were selected based on a review of published studies [3 (link),12 (link),30 (link)], recommendations from clinical co-investigators with expertise in fracture ascertainment in administrative databases (WDL, SM), and the authors’ previous experience ascertaining other chronic diseases, include osteoporosis, in administrative data [31 (link)-33 (link)]. The case definitions were differentiated by: (a) source of data, (b) number of records with the relevant diagnosis code(s), (c) type of diagnosis in hospital data, (d) presence of service codes in physician billing claims, and (e) duration of the fracture-free period. With the exception of one hip fracture case definition, all site-specific definitions used the same ICD-9-CM and ICD-10-CA diagnosis code(s). For hip fracture, we considered ICD-9-CM 820 (fracture of neck of the femur) and 821 (fracture and other unspecified parts of the femur) because some hip fractures may be assigned a less precise diagnosis code [34 (link)]. Case definitions were based on hospital data only (hip) or hospital and physician claims data, in keeping with previous research [3 (link),15 (link)]. For the latter, case definitions requiring one or at least two records with the specified diagnosis code(s) were considered. Service codes capture radiologic and magnetic resonance imaging services for incident clinical vertebral fracture, immobilization or fixation services for wrist fracture, and surgical repair and fixation procedures for hip fracture. Service codes have also been used in previous studies to improve fracture ascertainment [35 (link)]. Fracture-free periods of zero, six or twelve months were considered, using the site-specific fracture index date to establish the end-point of the fracture-free period.
To illustrate the interpretation of the case definitions, H1 identifies hip fractures using hospital records with ICD-9-CM 820 or 821 (ICD-10-CA S72.0, S72.1, or S72.2) as the most responsible (i.e., primary) diagnosis; it does not use physician service codes nor does it require a fracture-free period. In contrast, case definition H13 identifies hip fractures from hospital records with ICD-9-CM 820 (ICD-10-CA S72.0, S72.1, or S72.2) in any diagnosis field. A physician service code was present within the hospitalization period and a 12-month fracture-free period was adopted. For wrist fracture, case definition W1 identifies fractures using hospital or physician billing records with ICD-9-CM 813 (ICD-10-CA S52) in any diagnosis field. This case definition requires a physician service code to accompany the diagnosis code and does not adopt a fracture-free period.
The fracture index date was the date of the first diagnosis or service code for a fracture event. Pathologic fractures were included because they represent a small proportion of all fractures and their exclusion can lead to underestimation of the fracture burden due to osteoporosis [36 (link)]. For each case definition, the number of incident fractures was generated for the Manitoba population 50years of age and older for fiscal years 1997/98 to 2006/07. Age, which was defined using the fracture index date, was obtained from health insurance registration files. For hip fracture, counts of incident fractures were generated both including and excluding residents of long-term care (i.e., nursing home) facilities [37 (link)]; the CaMos data excludes residents of these facilities and this may affect comparability of estimates. Residence in a facility was determined from nursing home files containing admission and separation dates.

The statistical package SAS was used for the analyses in this study. SAS was used to select the study and comparison groups (SAS Institute Inc., Cary, NC, USA). Descriptive analyses of the independent variables (patient characteristics, demographics, personal history at baseline, surgical interventions, the amount of time between fracture and the onset of depression and other comorbidities) are reported as percentages or the mean ± standard deviation (SD). The X² test was used to make between-groups comparisons of patient demographics, including economic status (monthly income: USD$>1000, USD$601∼1000, USD$<600), urbanization of their home city (level 1 to 4), the geographic location of patients’ residence (northern, central, southern, and eastern Taiwan), personal history at baseline (diabetes mellitus, hypertension, renal failure, liver cirrhosis, stroke and osteoporosis) and mortality rate between patients with fracture and non-fracture. The urbanization of patients’ home city was defined by population and certain indicators of the city’s level of development. Level 1 urbanization was defined as having a population greater than 1,250,000 people and a specific status of political, economic, cultural and metropolitan development. Level 2 urbanization was defined as having a population between 500,000 and 1,250,000 and an important role in the Taiwanese political system, economy and culture. Urbanization levels 3 and 4 were defined as having a population between 150,000 and 500,000 and less than 150,000, respectively. Furthermore, a crude hazard ratio (HR) was calculated using Cox’s stratified proportional hazards model (stratified with age, sex and the number of years since index hospitalization) to analyze the risk of new-onset major depression between the study and comparison groups. The covariate-adjusted HR was analyzed after adjusting for diabetes mellitus, hypertension, renal failure, liver cirrhosis, stroke, osteoporosis, geographic regions, post-fracture co-morbidities, monthly income and urbanization of patients’ home cities. In addition, we used SAS to analyze the eight most common post-fracture comorbidities during the study period. The relationship between post-fracture comorbidities and major depression was also analyzed. We used the Kaplan-Meier method and Log-rank test to estimate survival curves and compare the 3-year major depression-free survival rate between patients with femoral neck fracture and those without. Among the femoral neck fracture patients, the amount of time before the onset of major depression was recorded and divided into 6 periods (<200, 201–400, 401–600, 601–800, 801–1000 and >1000 days). Additionally, the relationship between surgical interventions (including hip replacement and open reduction of internal fixation of the hip) and the chance of suffering new-onset major depression was analyzed for this group (X² test). Hip replacement included hip arthroplasty in this study. A p-value <0.05 was considered to be statistically significant.

Human MSC of non-osteoporotic donors were obtained from bone marrow of femoral heads according to the described protocol [53] (link) after total hip arthroplasty due to osteoarthritis and/or hip dysplasia. MSC of patients suffering from osteoporosis were isolated from femoral heads after low-energy fracture of the femoral neck. Additional criteria for confirming primary osteoporosis in these donors were vertebrae fractures and advanced age.
Cell culture medium, fetal calf serum (FCS), trypsin-EDTA and antibiotics were obtained from PAA Laboratories GmbH, Linz, Austria. Human MSC were selected by surface adherence and expanded in DMEM/Ham's F-12 (1∶1) medium supplemented with 10% heat-inactivated FCS, 1 U/ml penicillin, 100 µg/ml streptomycin and 50 µg/ml L-ascorbic acid 2-phosphate (Sigma Aldrich GmbH, Schnelldorf, Germany).
For long term cultivation, cells were expanded at 70–90% confluence by trypsinization with 1× trypsin-EDTA and reseeding in a ratio of 1∶3. This procedure was repeated for up to x passages when the hMSC did not become confluent within 3 weeks due to replicative senescence.

Data used in this secondary analysis of 404 patients with acute HFs were from two randomized controlled trials (RCT1 and RCT2). Both RCT were performed in the orthopedic- and the geriatric departments of Umeå University Hospital, Sweden. RCT1 included 199 patients with femoral-neck fractures enrolled in May 2000-December 2002 [20 (link)]; RCT2 included 205 patients with femoral-neck and trochanteric-region fractures enrolled in May 2008-June 2011 [21 (link)]. Inclusion criteria for both studies were age ≥ 70 years and Umeå municipality residence. Exclusion criteria for RCT1 were severe rheumatoid arthritis/hip osteoarthritis, pathological/hospital-acquired HF, renal failure, and bedridden before the fracture occurred. Those for RCT2 were hospital-acquired/pathological HF. Both RCTs included patients with cognitive impairment or dementia.

The clinical bone tissue samples of the experimental group used in this paper are from 15 patients with steroid induced avascular necrosis of the femoral head, including nine male patients and six female patients. Another 15 femoral head tissue samples from patients undergoing total hip replacement (The artificial prosthesis, including the femur and acetabulum, is fixed on the normal bone by bone cement and screws to replace the diseased joint) due to femoral neck fracture are used as the control group in this article, including eight male and seven female patients with femoral neck fracture. All patients participating in the study signs the informed consent form, and obtains detailed clinical pathology and prognosis data of all patients in the experimental group and the control group. The study covering relevant patient screening, clinical examination, clinical data acquisition and subsequent experimental analysis. The collected femoral head bone tissue samples are frozen in the prepared liquid nitrogen immediately after the operation for subsequent experimental analysis of total RNA and protein sample extraction.
Cell line: The product classification numbers of hFOB1.19 and human renal epithelial cell line 293 T are CRL-11372 and CRL-3216 respectively.
Main reagents and antibodies: conventional chemical reagents: disodium hydrogen phosphate, methanol, anhydrous ethanol, glacial acetic acid, isopropanol and other conventional reagents used in this paper are analytical pure. Total RNA extraction reagent (Trizol solution) is purchased from Bioengineering Co., Ltd. (Shanghai), and the product number is B511311. Diethyl pyrocarbonate is purchased from Sigma Aldrich (China) Co., Ltd. (Shanghai, China), and the product number is 472565. Agarose is purchased from Yisheng Biotechnology Co., Ltd. (Shanghai, China), and the product number is 10208ES60.
DNA marker is purchased from Beyotime Biotechnology Company (Nantong, China), and the product number is D0107. Nucleotide fluorescent dye (SYBR GreenI) is purchased from Thermo Fisher Scientific (China) Co., Ltd. (Shanghai, China), and the product number is S7563. Taq DNA polymerase is purchased from Fullgold Biotechnology Co., Ltd. (Beijing, China), and the product number is AP101-12. The basic idea of clinical experiment is shown in Figure 2.

Cell Counting Kit-8 (CCK8) method is used to detect the cell viability of cultured human hFOB1. 19 osteoblasts. The experimental operation process is carried out according to the technical methods provided in the manual. The simple experimental process is as follows. After transfection, hFOBl. 19 osteoblasts are cultured for 24–27 h and used for CCK-8 analysis of cell activity. Cells in each group are collected at three different time points, namely, 24, 48 and 72 h. First, pipettes are used to gently remove old media. Then, according to the ratio of 100:1, the fresh complete culture medium is diluted with CCK-8 reagent. Finally, the cell culture plate $\mathrm{n}\left(\mathrm{j}\right)$ is put back into the incubator to continue to culture for about 1 h, and then detected by the microplate reader for statistical analysis of the difference ${\mathrm{c}}_{\mathrm{y}}$ in cell activity among groups. At least three biological repeats shall be set for the above analysis (Wang et al., 2019 (link)). $k_g=g_{FS}-g$
$\mathrm{n}\left(\mathrm{j}\right)=\frac{∆\mathrm{l}}{∆\mathrm{z}*\mathrm{n}}$
${\mathrm{a}}_{\mathrm{a}\mathrm{s}}=\frac{1}{\mathrm{d}\left(\mathrm{\lambda }\right)}$
${\mathrm{c}}_{\mathrm{y}}=\mathrm{h}+\frac{\mathrm{g}}{{\mathrm{P}}_{\max }\bullet {\mathrm{t}}_0}$
During the miR-206 expression level difference experiment, the femoral head tissue samples of 15 patients with femoral neck fracture who underwent total hip arthroplasty were collected as the control group.