【Abstract】Objective To observe the bacteria in cholesterol stones by electronic microscope and to explore the role of bacteria in the stone formation.Methods Twelve patients with cholelithiasis underwent operations (male 6, female 6, average age 54.6 years） with cholecystolithiasis 5, extrahepatic and intrahepatic bile duct stone 1, common bile duct stone combining with gallstone 6. The cholesterol stones were observed by electronic microscope.Results There were bacterial structures in the cholesterol stones and cholesterol crystals.Conclusion There are bacteria in the core and peripheral of cholesterol stones, which suggests that bacteria may play an initial role in the formation of cholesterol stones.
Objective To comprehend the pathological features and possible pathogenesis of avascular necrosis of the femoral head (ANFH) by morphology and immunohistochemical observation of osterix (OSX) and adiponectin through in vitro traumatic and non-traumatic ANFH specimens, so as to provide a theoretical basis for cl inical treatment. Methods Sixty-six ANFH specimens were collected from 66 cl inical cases undergoing hip replacement surgery. Twenty-four cases of traumatic ANFH (group A) included 17 males and 7 females, aged 21 to 70 years with an average of 56.5 years; 23 cases of steroid-induced ANFH (group B) included 16 males and 7 females, aged 56 to 72 years with an average of 61 years; and 19 cases of alcohol ic ANFH (group C) were males, aged 55 to 67 years with an average of 58.5 years. Bone tissue was got from weight-bearing and non-weight-bearing area of the femoral head respectively. The basic pathological changes was observed by HE staining under the optical microscope, and the percentage of empty bone lacuna and the percentage of trabecular bone area were calculated. The morphological changes of ANFH in different groups were observedby scanning electron microscope (SEM). OSX and adiponectin expression were detected by immunohistochemical technique. Results Gross of the femoral head surface in each group was rough, collapse, articular cartilage loss, osteophyte formation; cross section: dark red in group A, and yellow in groups B and C. HE staining showed that weight-bearing area of ANFH have similar morphological features in three groups. In non-weight-bearing area of groups B and C, the fat cells in bone marrow markedly increased and were hypertrophic; however there were more fibrous tissue in group A. There were statistically significant differences (P lt; 0.001) in the percentage of empty bone lacuna of the weight-bearing and non-weight-bearing area among three groups. There were no statistically significant differences (P gt; 0.05) in the percentage of trabecular bone area among three groups. The SEM observation showed that three groups had similar pathological changes. Brown granules for OSX and adiponectin positive substance were mainly located in the osteoblast of bone marrow of the femoral head. There was statistically significant difference (P lt; 0.05) in the average absorbency (A) value of OSX between group A and groups B, C, but there was no statistically significant difference (P gt; 0.05) between groups B and C. While there was no statistically significant difference (P gt; 0.05) in the A value of adiponectin among three groups. Conclusion Hormones and alcohol necrosis have more obviously fatty degeneration, but the repair capacity of traumatic femoral head necrosis is ber than that of hormones and alcohol necrosis. Alcohol and hormones have inhibitory action on the OSX-mediated osteogenic differentiation. Hormones and alcohol may not affect osteoblast expressing adiponectin and its receptors.
The development of intravital microscopy (IVM) has enabled researchers to perform in-situ, real-time observations of pulmonary micro-circulation at the cellular level, and has become instrumental for researching the immune micro-environment of pulmonary diseases. This article introduces the developments in constructing the pulmonary imaging window and summarizes the current light microscopy techniques used for lung IVM with regard to its relevant functional and application features, which includes wide field fluorescence microscopy, confocal microscopy, as well as two-photon microscopy. It then provides examples of IVM of pulmonary immune response in inflammation and infection in murine models, and finally specifies the technological limitations to provide reference for researchers to systematically learn and understand the technology.
Objective To investigate the feature and regularity of the collagen change in bone healing during bone lengthening. Methods Bone lengthening model was made in the middle segment of the rabbit tibia. Five days after the model was established, the bone was lengthened 1.5 mm perday for 14 days. The rabbits were put to death after elongation, 7,14,21,30,40,50,60 and 70 days after elongation. The distracted area of the bone was imbedded with paraffin. After being stained by the picric acidsirius red staining, the slice was observed under polarized microscope. Results The features of the collagen change in the distracted bone were as follows: ① In the fibrous tissue of the distracted area during lengthening period and the early stage after lengthening, there was not only collagen Ⅲ but alsomuch collagen Ⅰ. ② Collagen Ⅰ, Ⅱ and Ⅲ were observed in the cartilage. ③ Collagen Ⅰ, Ⅱ and Ⅲ were also observed in the pseudogrowth plate. ④ Collagen Ⅰ took the dominance during lengtheningperiod and the late stage after lengthening. Conclusion New bone formation in bone lengthening is under the distracted force, so the collagen changes have different features compared with that in fracture healing. Collagen Ⅰ, Ⅱ and Ⅲcan be identified by picric-acid-sirius red staining and polarized microscope, so a new method for studying the collagen typing in bone repairing is provided.
Objective To investigate the microscope-assisted anterior cervical surgery and traditional open surgery for the treatment of cervical myelopathy with ossification of the posterior longitudinal ligament (OPLL). Methods Retrospective selection of patients with OPLL who underwent microscope-assisted and traditional open anterior cervical surgery in West China (Airport) Hospital Sichuan University were selected between January 2016 and August 2020. The patients who underwent traditional open anterior cervical surgery between January 2016 and August 2018 were classified as the conventional group, and the patients who underwent microscope-assisted anterior cervical surgery between September 2018 and August 2020 were classified as the microscope group. The baseline characteristics, operative time, intraoperative blood loss, length of hospital stay, Visual Analogue Scale (VAS) of pain before and after surgery, and surgical complications were collected. Neurological function was assessed using the Japanese Orthopaedic Association (JOA) score. Result A total of 46 patients were included. There were 24 cases in the conventional group and 22 cases in the microscope group. There was no significant difference in baseline characteristics between the two groups (P>0.05). The operation time, intraoperative blood loss and length of hospital stay in the microscope group were lower than those in the conventional group (P<0.001). There was no significant difference in VSA score and JOA score between the two groups before operation (P>0.05). There were statistically significant differences in VAS score and JOA score between the two groups 18 months after operation (P<0.001). The comparison of VAS score and JOA score in the two groups before and after operation showed that there was a statistically significant difference between 18 months after operation and before operation (P<0.05). In the microscope group, the average improvement rate of neurological function [(79.90±16.67)% vs. (58.12±17.47)%, t=4.317, P<0.001], excellent and good rate [95.45% (21/22) vs. 66.67% (16/24), χ2=4.354, P=0.037] were higher than those in the conventional group. The total number of complications in the microscope group was lower than that in the conventional group (P=0.024). Conclusion Compared with the traditional open anterior cervical surgery, the microscope-assisted anterior cervical surgery for OPLL can reduce intraoperative blood loss and length of hospital stay, reduce the incidence of postoperative complications.
Objective To investigate the assembl ing and cl inical appl ication of the video output system util izing teaching sight glass of surgical microscope. Methods Between June 2009 and April 2010, 10 patients with craniocervical junction malformation were treated by the method of transoral-transpharyngeal approach with the microscope and videooutput system under the direct vision. There were 6 males and 4 females with an average age of 32 years (range, 13-52 years). Three cases had the history of injury and 7 cases had no history of definite injury. The disease duration was from 10 months to 12 years (median, 5 years). The main cl inical symptoms were brevicoll is or torticoll is; 2 patients had malformation appearance and 4 patients had occi put-cervical pain. The physical examination showed that all patients had the symptoms that upper cervical cord was damaged; the imaging examination showed that all patients had basilar invagination, atlantoaxial dislocation, and ossification. Before and after operations, the functions of nerve were evaluated by Japanese Orthopaedic Association (JOA) scoring, the improvement rate was calculated to evaluate the efficacy. Results By the video output system assembly, 15.1 mill ion pixels high-definition images could be collected and reached 1 920 × 1 080 pixels video camera, so assistants or medical students could watch the cl inical operation directly. All patients had no neural injury or cerebrospinal fluid leakage during operation. Basilar invagination and atlantoaxial dislocation were corrected. Infection at incision occurred in 1 patient; other incisions healed by first intention without early compl ication. All patients were followed up 6-16 months (mean, 13.5 months). The average JOA score was increased from 10.2 preoperatively to 15.5 at 6 months postoperatively with an improvement rate of 77.9%. At 12 months after operation, bony fusions were achieved. Conclusion The miscroscope and video output system can improve the effectiveness of the original surgical microscope. It makes visual fields much clearer and operations more accuratewith a few compl ications.
Objective The combined appl ication of green fluorescent protein (GFP) and confocal laser scanning microscope three-dimensional reconstruction (CLSM-3DR) were used to monitor the construction and in vivo transplantation of tissue engineered bone (TEB), to provide for technology in selection of scaffolds and three-dimensional constructional methods. Methods After bone marrow mesenchymal stem cells (BMSCs) were isolated from a 2-year-old green goat by a combination method of density gradient centrifugation and adherent culture, and the expressions of CD29, CD60L, CD45, and CD44 in BMSCs were detected by flow cytometry. Plasmid of pLEGFP-N1 was ampl ified, digested by enzymes (Hind III, BamH I, Sal I, and Bgl II), and identified. Transfection of pLEGFP-N1 into PT67 cells was performed under the help of l iposome. Positive PT67 cells were picked out with G418, and prol iferated for harvesting virus. Based on the titre of virus, after BMSCs were infected by virus containing pLEGFP-N1, GFP positive BMSCs were collected and prol iferated for seeding cells. TEB was fabricated by GFP positive BMSCs and decalcified bone matrix (DBM) and observed by CLSM-3DR for the evaluation of the distribution and prol iferation of seeding cells. After TEB was transplanted in the defect of goat femur, CLSM was used for observing the survival and distribution of GFP positive cells in the grafts. Results The isolated cells were fibroblast-l ike morphous, with the positive expression of CD29 and CD44, and negative expression of CD60L and CD45. The digested production of pLEGFP-N1 was collected for ionophoresis, whose results showed the correct fragment length (6 900 bp). The virus of pLEGFP-N1 was harvested by transfection of pLEGFP-N1 into PT67 cells and used for further infection to obtain GFP positive BMSCs. The prol iferated GFP positive BMSCs and DBM were used for fabrication of TEB. The distribution, prol iferation, and migration of BMSCs in TEB were observed by CLSM-3DR. GFP positive cells also were observed in images of TEB graft in goat femur 28 days after transplantation. Conclusion The BMSCs labeled by GFP in three-dimensional scaffold in vivo were monitored well by CLSM-3DR. It suggests a wide use potency in monitoring of three-dimensional cultured TEB.
OBJECTIVE: To investigate the expression and distribution of platelet derived growth factor receptor-beta(PDGFR-beta) in normal skin and keloid and to discuss its biological function in keloid formation. METHODS: 1. To detect the expression and distribution of PDGFR-beta in normal skin and keloid tissue by immunohistochemistry; 2. To detect the receptor expression in vitro by Flow cytometry (FCM); 3. To detect the subcellular distribution of receptor by Laser confocal microscope. RESULTS: 1. Immunohistochemistry showed that normal skin and keloid tissue were almost the same in expression but different in distribution of PDGFR-beta; 2. There was more expression of PDGFR-beta in normal fibroblasts than that in keloid fibroblasts in vitro by FCM; 3. Laser confocal microscope revealed that the PDGFR-beta concentrated on the surface of cell membrane in keloid fibroblasts, but in normal skin fibroblasts, the receptors were coagulated on the nuclear membrane and intranucleus. CONCLUSION: Compared with the fibroblasts in vivo, there was a difference of the PDGFR-beta expression in fibroblasts in vitro, more expression of PDGFR-beta in normal fibroblast than that in keloid fibroblast in vitro; and the subcellular distribution of PDGFR-beta was different in normal skin and keloid fibroblasts. The characteristics of the expression and distribution of PDGFR-beta in keloid may contribute to the formation of keloid.
Objective To study the effect of substance P ( SP) on int racellular f ree calcium concent ration in human poorly-differentiated gast ric cancer cell in vitro. Methods Human gast ric cancer cell line MKN45 was cultured in RPMI 1640. Then the cells were loaded with specific calcium fluorescent probe Furu23/ AM. ASN21377642 (NK21 receptor antagonist) , Nicardipine (calcium channel blocker) and different concent rations of SP were used to treat gast ric cancer cells. The concent ration changes of int racellular free calcium were detected by laser scanning confocal microscope. Results It was found that 10 , 50 and 100 nmol/ L SP could significantly increase the int racellular free calcium concent ration of gast ric cancer cells in Hanks solutions , which contain ext racellular calcium ( P lt;0. 05) , and the change was in a dose-dependent manner ( P lt; 0. 05) . When there was ext racellular calcium existed ,the increasing amplitude of intracellular f ree calcium concent ration was significantly higher than that when there was no extracellular calcium ( Plt; 0. 05) . And when Hanks solutions were pretreated with ASN21377642 and Nicardipine , the effects of 100 nmol/ L SP were partly inhibited , and the concent rations of int racellular f ree calcium were significantly lower than those in group s without pret reatment s ( P lt; 0. 05) . Conclusion SP can significantly increase free calcium concent ration in the gastric cancer cells. Releasing of stored calcium in the cells and influx of extracelluar calcium may contribute to the elevation of int racellular free calcium concentration.
Objective To explore the feasibility of microscopic spermatic vein ligation for unilateral varicocele under daytime surgical mode. Methods The clinical data of patients with unilateral varicocele who underwent daytime surgical treatment at the General Hospital of Northen Theater Command between January and December 2022 were retrospectively analyzed. The relevant indicators of patients were collected, including age, surgical time, intraoperative blood loss, total hospitalization expenses, and postoperative condition. Results A total of 159 patients were included, aged 18-41 years, with an average age of (27.79±4.80) years, an average surgical time of (49.23±5.57) minutes, and an average intraoperative blood loss of (8.94±3.58) mL. One patient had fever, nausea and acid regurgitation on the 2nd day after discharge, and one patient had urinary retention on the day after operation. Five patients had a pain score of ≥3 in the surgical area on the 1st day after surgery. All patients had no postoperative incisions with bleeding or hematoma. Follow up showed that all patients returned to normal life on the 3rd day after surgery and returned to work on the 10th day after surgery. At 3-6 months after surgery, the sperm quality of 94 patients improved. One patient experienced disease recurrence 6 months after surgery. Conclusion Microscopic spermatic vein ligation for unilateral varicocele under daytime surgical mode is safe and feasible, and is worthy of promotion.