lncRNA-KCNQ1OT1: A Potential Target in Exosomes Derived from Adipose-Derived Stem Cells for the Treatment of Osteoporosis.

BACKGROUND: Osteoporosis is a worldwide medical and socioeconomic burden characterized by systemic impairment of bone strength and microstructure. Exosomes derived from adipose-derived stem cells (ADSCs-Exos) have been confirmed to play effective roles in the repair of various tissues and organs. This study was aimed at investigating the role of ADSCs-Exos and a novel long noncoding RNA KCNQ1OT1 played in osteoporosis as well as the underlying mechanism. METHODS: Primary osteoblasts were treated with different doses of tumor necrosis factor-α (TNF-α) (0, 1, 2.5, 5, and 10 ng/ml) and then cocultured with ADSCs-Exos or exosome-derived from lnc-KCNQ1OT1-modified ADSCs (KCNQ1OT1-Exos). The expression of miRNA-141-5p (miR-141-5p) and lnc-KCNQ1OT1 was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). The protein expression of cleaved-caspase-3, caspase-3, and Bax was determined by Western blot. Cell viability and apoptosis were assessed by Cell Counting Kit-8 (CCK-8) and flow cytometry analysis, respectively. The binding sites between KCNQ1OT1 and miR-141-5p were validated by dual-luciferase reporter assay. RESULTS: TNF-α dose-dependently increased miR-141-5p expression, inhibited viability, and promoted apoptosis of osteoblasts. However, miR-141-5p silencing or cocultured with ADSCs-Exos attenuated these effects. In addition, KCNQ1OT1-Exos could more significantly attenuate the induced cytotoxicity and apoptosis compared to ADSCs-Exos. Moreover, miR-141-5p was confirmed as the target of KCNQ1OT1 by luciferase reporter assay. CONCLUSIONS: ADSCs-Exos can attenuate cytotoxicity and apoptosis of TNF-α-induced primary osteoblasts. KCNQ1OT1-Exos have a more significant inhibitory effect compared to ADSCs-Exos by the function of sponging miR-141-5p, suggesting that KCNQ1OT1-Exos can be promising agents in osteoporosis treatment.

[Intra-articular injection of compound betamethasone and hyaluronic acid for the treatment of moderate-severe knee osteoarthritis:a randomized controlled trial].

OBJECTIVE: To compare clinical effects of compound betamethasone and compound betamethasone with hyaluronic acid in treating moderate-severe knee osteoarthritis (KOA). METHODS: A prospective randomized controlled study was conducted in 116 patients with unilateral moderate-severe KOA patients from February 2017 to November 2017 and divided into observation group and control group, 58 patients in each group. In observation group, there were 15 males and 43 females aged from 45 to 80 years old with an average of (66.45±6.31) years old;according to Kellgren-Lawrence(K-L) classification, 42 patients were type Ⅲ and 16 patients were type Ⅳ;the courses of disease ranged from 4 to 8 years with an average of (5.25±2.21) years;the patients were treated by injecting 1 ml compound betamethasone into knee joint. In control group, there were 13 males and 45 females aged from 45 to 80 years old with an average of (64.89±6.41) years old;according to K-L classification, 43 patients were type Ⅲ and 15 patients were type Ⅳ;the courses of disease ranged from 4 to 10 years with an average of (5.41±2.35) years;the patients were treated by knee joint injection of 4 ml hyaluronic acid and 1 ml compound betamethasone. Visual analog scale (VAS), Western Ontario and McMaster University Osteoarthritis Index (WOMAC) were used to evaluate clinical effects before treatment and 1 week, 1, month, 3 and 6 months after treatment. RESULTS: Totally 55 patients in observation group were followed up for 6 months, and 3 patients were quit at 3 months after treatment for poor efficacy. Totally 56 patients in control group were followed up for 6 months, and 2 patients were withdrew from the follow-up on the first and third month respectively for poor efficacy. There were no statistical difference in VAS and WOMAC between two groups before treatment and different time points after treatment (P>0.05), and no significant difference in VAS and WOMAC before treatment and 6 months after treatment between two groups(P>0.05). CONCLUSION: For patients with moderate-severe KOA, there is no significant difference in therapeutic effect between compound betamethasone injection and compound betamethasone combined with hyaluronic acid injection, and long-term effect of two methods is not good.

[Study on pain control of continuous adductor block analgesia after primary knee replacement].

OBJECTIVE: To investigate the effect of continuous adductor block on pain control after bilateral knee joint Ⅰ stage replacement. METHODS: A retrospective analysis was made of the data of 24 patients with bilateral knee joint I stage replacement who were treated in our hospital from January 2018 to January 2019, and who underwent continuous adductor block analgesia. There were 6 males and 18 females, aged 60 to 72 (65.05±5.82) years old. The patients underwent continuous block of adductor canal with patient-controlled analgesia system. At 4, 6, 12, 24, 36 and 48 hours after operation, visual analogue score(VAS) of resting state and passive motion state was performed;the knee joint activity was followed up for 1 week, 1, 3 and 6 months after operation;the knee joint function was scored at 6 months after operation, using the knee joint scoring standard of American Special Surgery Hospital(HSS);adverse reactions and complications were recorded. RESULTS: The VAS scores under resting state and passive motion state at each time point were less than 3 points in patients with continuous adductor block. The patients had better postoperative exercise of knee joint activity. The score of HSS was excellent in 20 cases, good in 2 cases, fair in 1 case and poor in 1 case. There were only 4 cases of nausea and vomiting, none of them had serious adverse reactions and complications such as bradycardia and deep vein thrombosis. CONCLUSION: Continuous adductor block has a significant effect on pain control and less adverse reactions after bilateral knee jointⅠ -stage replacement.

The Differential Effects of Leukocyte-Containing and Pure Platelet-Rich Plasma on Nucleus Pulposus-Derived Mesenchymal Stem Cells: Implications for the Clinical Treatment of Intervertebral Disc Degeneration.

BACKGROUND: Platelet-rich plasma (PRP) is a promising strategy for intervertebral disc degeneration. However, the potential harmful effects of leukocytes in PRP on nucleus pulposus-derived mesenchymal stem cells (NPMSCs) have seldom been studied. This study aimed at comparatively evaluating effects of pure platelet-rich plasma (P-PRP) and leukocyte-containing platelet-rich plasma (L-PRP) on rabbit NPMSCs in vitro. METHODS: NPMSCs isolated from rabbit NP tissues were treated with L-PRP or P-PRP in vitro, and then cell proliferation and expression of stem cell markers, proinflammatory cytokines (TNF-α, IL-1ß), production of ECM (extracellular matrix-related protein), and NF-κB p65 protein were validated by CCK-8 assay, real-time polymerase chain reaction, enzyme-linked immunosorbent assay, immunofluorescence, and western blot respectively. RESULTS: NPMSCs differentiate into nucleus pulposus-like cells after treatment of PRPs (P-PRP and L-PRP), and NPMSCs exhibited maximum proliferation at a 10% PRP dose. L-PRP had observably higher concentration of leukocytes, TNF-α, and IL-1ß than P-PRP. Furthermore, compared to P-PRP, L-PRP induced the differentiated NPMSCs to upregulate the expression of TNF-α and IL-1ß, enhanced activation of the NF-κB pathway, increased the expression of MMP-1 and MMP-13, and produced less ECM in differentiated NPMSCs. CONCLUSIONS: Both P-PRP and L-PRP can induce the proliferation and NP-differentiation of NPMSCs. Compared to L-PRP, P-PRP can avoid the activation of the NF-κB pathway, thus reducing the inflammatory and catabolic responses.

Intra-articular, single-shot co-injection of hyaluronic acid and corticosteroids in knee osteoarthritis: A randomized controlled trial.

The aim of the present study was to investigate whether the co-injection of hyaluronic acid (HA) and corticosteroids (CS) was superior to HA alone in the treatment of knee OA. A total of 120 participants with symptomatic knee OA were recruited and formed the intention-to-treat population for a 6-month follow-up. In the HA group, patients received a single-shot injection of 4 ml HA. In the HA&CS group, patients received a co-injection of 3 ml compound betamethasone solution and 4 ml HA. Visual analog scale (VAS), Western Ontario and McMaster University Osteoarthritis Index (WOMAC) and knee flexion motion were assessed as primary outcomes. Patients in the HA&CS group exhibited better pain relief and knee function at the time points of week 1, month 1 and month 3 (P<0.05). For the last follow-up at month 6, the values did not differ significantly between these two groups. Patients in both groups exhibited improvement in pain, knee function, and range of motion following injection. For the final follow-up at month 6, the mean VAS score, WOMAC score and knee flexion motion were still superior to that prior to treatment, but the values did not differ significantly. The co-injection of HA and CS provided a rapid improvement in pain relief, knee function, and range of motion, but did not differ significantly from that of HA alone in the long term effect.

Is exclusion of leukocytes from platelet-rich plasma (PRP) a better choice for early intervertebral disc regeneration?

BACKGROUND: Platelet-rich plasma (PRP) is becoming a promising strategy to treat early intervertebral disc degeneration (IDD) in clinics. Pure PRP without leukocytes (P-PRP) may decrease the catabolic and inflammatory changes in the early degenerated intervertebral discs. The aim of this study was to investigate the effects of P-PRP on nucleus pulposus-derived stem cells (NPSCs) isolated from early degenerated intervertebral discs in vitro. METHODS: NPSCs isolated from early degenerated discs of rabbits were treated with P-PRP or leukocyte-platelet-rich PRP (L-PRP) in vitro, followed by measuring cell proliferation, stem cell marker expression, inflammatory gene expression, and anabolic and catabolic protein expression by immunostaining, quantitative real-time polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay. RESULTS: Cell proliferation was induced by P-PRP in a dose-dependent manner with maximum proliferation at 10% P-PRP dose. P-PRP induced differentiation of NPSCs into active nucleus pulposus cells. P-PRP mainly increased the expression of anabolic genes and relative proteins, aggrecan (AGC), collagen types II (Col II), while L-PRP predominantly increased the expression of catabolic and inflammatory genes, matrix metalloproteinase-1 (MMP-1), MMP-13, interleukin-1 beta (IL-1ß), IL-6, tumor necrosis factor alpha (TNF-α), and protein production of IL-1ß and TNF-α. CONCLUSIONS: Leukocytes in PRP activate inflammatory and catabolic effects on NPSCs from early degenerated intervertebral discs. Hence, P-PRP may be a more suitable therapeutic strategy for early IDD.

Stem cell-derived exosomes: A promising strategy for fracture healing.

OBJECTIVES: To describe the biological characteristics of exosomes and to summarize the current status of stem cell-derived exosomes on fracture healing. Meanwhile, future challenges, limitations and perspectives are also discussed. METHODS: Search and analyze the related articles in pubmed database through the multi-combination of keywords like "stem cells","exosomes","bone regeneration" and "fracture healing". CONCLUSION: Stem cell-derived exosome therapy for fracture healing has been enjoying popularity and is drawing increasing attention. This strategy helps to promote proliferation and migration of cells, as well as osteogenesis and angiogenesis, in the process of bone formation. Although the exact mechanisms remain elusive, exosomal miRNAs seem to play vital roles. Future studies are required to solve multiple problems before clinical application, including comprehensive and thorough understanding of exosomes, the exact roles of exosomes in regulating bone formation, and the optimal source, dose and frequency of treatment, as well as technical and safety issues. Moreover, studies based on fracture models of large animals are could offer guidance and are in demand.

Intervertebral disc regeneration using platelet­rich plasma­containing bone marrow­derived mesenchymal stem cells: A preliminary investigation.

Platelet­rich plasma (PRP) is a promising strategy for intervertebral disc degeneration (IDD). However, the short half­life of growth factors released from PRP cannot continuously stimulate the degenerated discs. Thus, the present study hypothesized that the combined use of PRP and bone marrow­derived mesenchymal stem cells (BMSCs) may repair the early degenerated discs in the long term for their synergistic reparative effect. In the present study, following the induction of early IDD by annular puncture in rabbits, PRP was prepared and mixed with BMSCs (PRP­BMSC group) for injection into the early degenerated discs. As controls, phosphate­buffered saline (PBS; PBS group) and PRP (PRP group) were similarly injected. Rabbits without any intervention served as a control group. At 8 weeks following treatment, histological changes of the injected discs were assessed. Magnetic resonance imaging (MRI) was used to detect the T2­weighted signal intensity of the targeted discs at weeks 1, 2 and 8 following treatment. Annular puncture resulted in disc narrowing and decreased T2­weighted signal intensity. At weeks 1 and 3, MRI examinations showed regenerative changes in the PRP­BMSC group and PRP group, whereas the PBS group exhibited a continuous degenerative process of the discs. At 8 weeks post­injection, the PRP­BMSCs induced a statistically significant restoration of discs, as shown by MRI (PRP­BMSCs, vs.PRP and PBS; P<0.05), which was also confirmed by histological evaluations. Thus, compared with PRP, the administration of PRP­containing BMSCs resulted in a superior regenerative effect on the early degenerated discs, which may be a promising therapeutic strategy for the restoration of early degenerated discs.

Stem cell therapy: a promising biological strategy for tendon-bone healing after anterior cruciate ligament reconstruction.

Tendon-bone healing after anterior cruciate ligament (ACL) reconstruction is a complex process, impacting significantly on patients' prognosis. Natural tendon-bone healing usually results in fibrous scar tissue, which is of inferior quality compared to native attachment. In addition, the early formed fibrous attachment after surgery is often not reliable to support functional rehabilitation, which may lead to graft failure or unsatisfied function of the knee joint. Thus, strategies to promote tendon-bone healing are crucial for prompt and satisfactory functional recovery. Recently, a variety of biological approaches, including active substances, gene transfer, tissue engineering and stem cells, have been proposed and applied to enhance tendon-bone healing. Among these, stem cell therapy has been shown to have promising prospects and draws increasing attention. From commonly investigated bone marrow-derived mesenchymal stem cells (bMSCs) to emerging ACL-derived CD34+ stem cells, multiple stem cell types have been proven to be effective in accelerating tendon-bone healing. This review describes the current understanding of tendon-bone healing and summarizes the current status of related stem cell therapy. Future limitations and perspectives are also discussed.

Dysregulated miR-98 Contributes to Extracellular Matrix Degradation by Targeting IL-6/STAT3 Signaling Pathway in Human Intervertebral Disc Degeneration.

Intervertebral disc degeneration (IDD) is associated with dysregulated expression of microRNAs (miRNAs). However, the precise molecular mechanisms underlying this disorder remain unclear. Therefore, we tested the hypothesis that miRNAs modulate IDD through effects on the IL-6/STAT3 signaling pathway, a potential regulator of IDD. The miRNA expression profile was determined in nucleus pulposus (NP) tissues from patients with IDD and controls, employing miRNA microarray and quantitative real-time PCR (RT-qPCR). Biological functions of differential expression miRNAs were further investigated using immunofluorescent staining. Luciferase reporter assays and Western blotting were performed to determine miRNA targets. We identified 41 miRNAs that were differentially expressed in patients compared with controls. Following RT-qPCR confirmation, miR-98 was significantly downregulated in degenerative NP tissues. Moreover, its level was inversely correlated with grade of disc degeneration. Through gain-of-function and loss-of-function studies, miR-98 was shown to significantly promote type II collagen expression in NP cells. Interleukin-6 (IL-6) was identified as a target of miR-98. Knockdown of IL-6 induced effects on NP cells similar to those induced by miR-98. In contrast, IL-6 treatment abrogated the effects induced by miR-98 upregulation. Moreover, miR-98 dramatically suppressed expression of STAT3 target gene, MMP2. IL-6 treatment antagonized this effect, whereas knockdown of IL-6 by IL-6 short hairpin RNA (shIL-6) induced inhibitory effects on the expression of p-STAT3 and its main target genes, similar to miR-98. The mRNA level of IL-6 was inversely correlated with that of miR-98 in degenerative NP tissues. These results suggest the downregulation of miR-98 could promote IDD through the IL-6/STAT3 signaling pathway. Our findings also highlight miR-98 as a novel hopeful therapeutic target for IDD.

Downregulation of microRNA-193a-3p is involved in invertebral disc degeneration by targeting MMP14.

UNLABELLED: Accumulating evidence suggests that microRNAs (miRNAs) play an important role in intervertebral disc degeneration (IDD), but the precise role of specific miRNAs involved in this disease remains elusive. The purpose of this study was to identify IDD-specific miRNAs, followed by functional validation of results. MiRNA expression profile was determined in nucleus pulposus (NP) tissues from patients with IDD and controls, employing Solexa sequencing and quantitative real-time PCR (qRT-PCR). Biological functions of differential expression miRNAs were further investigated in vitro and in vivo. Luciferase reporter assays and Western blotting were performed to determine miRNA targets. We identified 28 miRNAs that were differentially expressed in patients compared with controls. Following qRT-PCR confirmation, miR-193a-3p was significantly down-regulated in degenerative NP tissues. Moreover, its level was correlated with grade of disc degeneration. Through gain- and loss-of-function studies, miR-193a-3p was demonstrated to significantly promote type II collagen expression in NP cells. Knockdown of MMP14 induced effects on NP cells similar to those induced by miR-193a-3p. Bioinformatics target prediction identified MMP14 as a putative target of miR-193a-3p. Furthermore, luciferase reporter assays and Western blotting demonstrated that miR-193a-3p directly targets MMP14. MiR-193a-3p inhibited IDD in vitro and in vivo. The downregulation of miR-193a-3p induces the expression of MMP14, which promotes loss of type II collagen and thereby contributes to the development of human IDD. Our findings extend the role of miR-193a-3p in the pathogenesis of IDD and provide a potential novel therapeutic target for degenerative disc disease. KEY MESSAGES: Intervertebral disc degeneration (ICC)-specific miRNA profile generated by next generation sequencing. Downregulation of miR-193a-3p promoted loss of type II collagen by directly targeting MMP14 in IDD. miR-193a-3p inhibited IDD in vitro and in vivo. miR-193a-3p may be a promising candidate for prevention of degenerative disc disease.

Effect of autologous platelet-rich plasma on the chondrogenic differentiation of rabbit adipose-derived stem cells in vitro.

This study aimed to isolate rabbit adipose-derived stem cells (ADSCs) and explore the potential of platelet-rich plasma (PRP) in the chondrogenic differentiation of ADSCs, thereby potentially providing a new approach for the repair and regeneration of cartilage injury. Rabbit ADSCs were isolated and characterized by induction towards adipogenic, osteogenic and chondrogenic lineages in vitro. The isolated ADSCs were also cultured with or without 10% PRP. Immunofluorescence staining, toluidine blue staining and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) were used to detect type II collagen (Col II) and aggrecan (AGC) expression. Col II immunofluorescence staining and toluidine blue staining indicated that following induction by autologous PRP, ADSCs manifested Col II and AGC expression. The expression of Col II and AGC mRNA was significantly upregulated in the PRP-treated cells when compared with that in control cells. Autologous PRP produced by laboratory centrifugation was able to promote the chondrogenic differentiation of rabbit ADSCs in vitro.

The Chondrogenic Induction Potential for Bone Marrow-Derived Stem Cells between Autologous Platelet-Rich Plasma and Common Chondrogenic Induction Agents: A Preliminary Comparative Study.

The interests in platelet-rich plasma (PRP) and their application in stem cell therapy have contributed to a better understanding of the basic biology of the prochondrogenesis effect on bone marrow-derived stem cells (BMSCs). We aimed at comparing the effect of autologous PRP with common chondrogenic induction agents (CCIAs) on the chondrogenic differentiation of BMSCs. Rabbit BMSCs were isolated and characterized by flow cytometry and differentiated towards adipocytes and osteoblasts. The chondrogenic response of BMSCs to autologous PRP and CCIAs which included transforming growth factor-ß1 (TGF-ß1), dexamethasone (DEX), and vitamin C (Vc) was examined by cell pellet culture. The isolated BMSCs after two passages highly expressed CD29 and CD44 but minimally expressed CD45. The osteogenic and adipogenic differentiation potentials of the isolated BMSCs were also confirmed. Compared with common CCIAs, autologous PRP significantly upregulated the chondrogenic related gene expression, including Col-2, AGC, and Sox-9. Osteogenic related gene expression, including Col-1 and OCN, was not of statistical significance between these two groups. Thus, our data shows that, compared with common chondrogenic induction agents, autologous PRP can be more effective in promoting the chondrogenesis of BMSCs.

Growth factors and platelet-rich plasma: promising biological strategies for early intervertebral disc degeneration.

Intervertebral disc degeneration (IDD) is a complex process with the mechanism not fully elucidated. The current clinical treatments for IDD are mainly focused on providing symptomatic relief without addressing the underlying cause of the IDD. Biological therapeutic strategies to repair and regenerate the degenerated discs are drawing more attention. Growth factor therapy is one of the biological strategies and holds promising prospects. As a promising bioactive substance, platelet-rich plasma (PRP) is considered to be an ideal growth factor "cocktail" for intervertebral disc (IVD) restoration. Results from many in vitro and in vivo studies have confirmed the efficacy of growth factors and PRP in IVD repair and regeneration. It is essential to advance the research on growth factor therapy and associated mechanism for IDD. This article reviews the background of IDD, current concepts in growth factor and PRP-related therapy for IDD. Future research perspectives and clinical directions are also discussed.

Enhancing intervertebral disc repair and regeneration through biology: platelet-rich plasma as an alternative strategy.

Intervertebral disc degeneration (IDD) is a common orthopedic disease associated with mechanical changes that may result in significant pain. Current treatments for IDD mainly depend on conservative therapies and spinal surgeries that are only able to relieve the symptoms but do not address the cause of the degeneration and even accelerate the degeneration of adjacent segments. This has prompted research to improve our understanding of the biology of intervertebral disc healing and into methods to enhance the regenerative process. Recently, biological therapies, including active substances, gene therapy and tissue engineering based on certain cells, have been attracting more attention in the field of intervertebral disc repair and regeneration. Early selection of suitable biological treatment is an ideal way to prevent or even reverse the progressive trend of IDD. Growth factors have been enjoying more popularity in the field of regeneration of IDD and many have been proved to be effective in reversing the degenerative trend of the intervertebral disc. Identification of these growth factors has led to strategies to deliver platelet-derived factors to the intervertebral disc for regeneration. Platelet-rich plasma (PRP) is the latest technique to be evaluated for promoting intervertebral disc healing. Activation of the PRP leads to the release of growth factors from the α-granules in the platelet cytoplasm. These growth factors have been associated with the initiation of a healing cascade that leads to cellular chemotaxis, angiogenesis, synthesis of collagen matrix, and cell proliferation. This review describes the current understanding of IDD and related biological therapeutic strategies, especially the promising prospects of PRP treatment. Future limitations and perspectives of PRP therapy for IDD are also discussed.

[Non-T-cell depleted HLA haploidentical peripheral blood stem cell transplantation for hematological malignancies: report of 36 cases].

OBJECTIVE: To analyze the clinical outcome of human leukocyte antigen (HLA) haploidentical peripheral blood stem cell transplantation (PBSCT) from related donors for hematological malignancies. METHODS: Thirty-six patients with hematological malignancies, with a median age of 25 (11-48) years, were transplanted with PBSC from an HLA-haploidentical family donors: 7 were 1 locus mismatched and 29 were 2-3 loci mismatched. The recipients received myeloablative conditioning regimen, in combination with different immunosuppressants according to the degree of HLA disparity followed by non-T-cell depleted PBSCT. The median number of CD34+ cells were 11 (4.16-21.00) x 10(6)/kg. RESULTS: All patients achieved sustained, full donor-type engraftment. Fifteen patients (41.7%) developed grade I-II aGVHD. Among 29 patients followed up more than 18 months, 17 (58.6%) developed cGVHD. There was no statistical difference in decrease and recovery of T, B and NK cell subsets after transplantation between HLA haploidentical group and HLA identical PBSCT group. The median follow-up duration was 15 (4 -69) months. Five patients (13.9% ) relapsed. The 2-year probability of leukemia-free survival (LFS) was 82.2%. CONCLUSION: Non-T-cell depleted HLA-haploidentical PBSCT is safe and feasible for patients with hematological malignancies after myeloablative conditioning regimen combined with intensive immunosuppressants.

[Establishment of obliterative bronchiolitis in allo-trachea transplant model of rat and detection of its pathogenesis preliminarily].

OBJECTIVE: To establish an animal model of obliterative bronchiolitis (OB) after lung transplantation and investigate the pathogenesis preliminarily. METHODS: Tracheal segments (5 cartilaginous rings each) were transplanted from SD rats to SD rats (Group I) or to Wistar rats (Group II and III). Grafts were implanted into an abdominal cavity and wrapped in the omentum. Animals in Group I and II did not receive CsA, animals in Group III received CsA daily by gastro-tube at 10 mg.kg(-1).d(-1) from beginning to end. Grafts were harvested on day 3, 14, 28 after transplantation as representative time points for 3 phases of injury in the evolution of allograft airway obliteration, then examined histological changes and gene expression of T-helper 1-and T-helper 2-type cytokines [Th1: interleukin-2 (IL-2), interferon-gamma (IFN-gamma); Th2: interleukin-4 (IL-4), interleukin-10 (IL-10)] in grafts. At the same time, effects of CsA were observed on the above-mentioned indices. RESULTS: There was no significant difference in histological changes on day 3 after transplantation among 3 groups (P > 0.05). Tracheas in Group I approached to normal morphology on day 14 after transplantation. Airway epithelium of Group II and III almost lost completely on day 14 after transplantation. There was no significant difference between Group II and Group III (P > 0.05), but there were significant differences between Group I and Group II or Group III. The cross-sectional area of the tracheal lumen was narrowed by approximately (5.0 +/- 1.2)%, (28.5 +/- 5.0)% and (19.4 +/- 2.9)% respectively on day 14 after transplantation in Group I, II and III, there were significant differences among 3 groups. On day 14 after transplantation, tracheas in Group I revealed few lymphocytic infiltration, but it showed dense lymphocytic infiltration in Group II. Tracheas in Group III have much more lymphocyte infiltration than that in Group I, but much less than that in Group II. There were significant differences among 3 groups, too (P < 0.01). The tracheal lumen revealed almost total luminal obstruction (94.8 +/- 3.6)% on day 28 after transplantation in Group II. The cross-sectional area of the tracheal lumen was narrowed by approximately (3.7 +/- 0.8)% and (36.6 +/- 7.6)% respectively in Group I and III on day 28. There were significant differences among 3 groups (P < 0.01). Compared with that on day 14, lymphocytic infiltration had decreased gradually on day 28 in Group II and III. There were significant differences among 3 groups all the same (P < 0.01). In Group II, expression of IL-2, IFN-gamma, IL-4, and IL-10 were much higher than that in Group I. Expression of Th1 cytokines was increased to a greater extent than that of Th2 cytokines in Group II compared with Group I. Allografts in Group III expressed significantly less IL-2 gene transcripts than that in Group II over all the points. There was no significant difference between Group II and III in IFN-gamma, IL-4, and IL-10 gene expression. CONCLUSIONS: Compared with isografts, allografts have more obvious changes, such as epithelial damage, fibroproliferation and lymphocytic infiltration. Th1 and Th2 lymphocyte subtypes contribute to the development of obliterative bronchiolitis in heterotopic trachea transplant model of rat, and changes of their cytokines gene expression may be involved in the pathogenesis. CsA could reduce the development of fibroproliferation and lymphocyte infiltration markedly, but it could not protect airway epithelium. CsA inhibits IL-2 gene transcripts, so it can reduce development of the pathologic lesion of obliterative bronchiolitis to a certain degree.

[Expression of CD80, CD86, TGF-beta1 and IL-10 mRNA in the esophageal carcinoma].

OBJECTIVE: To investigate the correlation of CD80 and CD86 mRNA expression with the expression of transforming growth factor-beta1 mRNA (TGF-beta1) and interleukin-10 mRNA (IL-10) in the esophageal cancer. To explore the reason of impaired immunological function of dentritic cell (DC) and the mechanism of cancer cell escaption from body immunity system in the esophageal cancer patient. METHODS: Expression of CD80, CD86, TGF-beta1 and IL-10R mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR) in specimens of 62 esophageal carcinoma and 16 normal esophageal mucosal tissues used as normal control. RESULTS: Expression of CD80 and CD86 mRNA in the esophageal cancer tissue was significantly lower than that in the normal esophageal mucosal tissue (CD80: P = 0.038; CD86: P = 0.0002). It was significantly higher in stage I or II than that in stage III or IV (CD80: P = 0.029; CD86: P = 0.045); and also higher in paitents with high or moderate differentiation than that with poor differentiation (CD80: P = 0.046; CD86: P = 0.044). Furthermore, it was found to be reversely correlated with expression of TGF-beta1, IL-10 mRNA by multiple regression analysis (P = 0. 0001) respectively, the more TGF-beta1 and IL-10 mRNA expressed in the tumor tissue, the less CD80 and CD86 mRNA expressed by dendritic cells. CONCLUSION: The expression of CD80 and CD86 mRNA in the tissues of esophageal cancer are found to be weak, and reversely correlated with the expression of TGF-beta1 and IL-10 mRNA. High level expression of TGF-beta1 and IL-10 mRNA may be an important influential factor to the weak expression of CD80 and CD86 mRNA, which may be one of the reasons leading to impaired function of dendritic cells and immune escape of cancer cells in the esophageal cancer patient.

[The expression of heparanase and vascular endothelial growth factor-C in human lung cancer].

OBJECTIVE: To investigate relationship between the expression of heparanase (HPSE) and vascular endothelial growth factor-C (VEGF-C) and tumorigenesis, progression in human lung cancer. METHODS: The expression of HPSE and VEGF-C protein in 65 cases of lung cancer, adjacent tissues of cancer and normal tissues was tested by immunohistochemical SABC method and analysed by clinico-pathological characteristics and prognosis of lung cancer. RESULTS: The rate of expression of HPSE and VEGF-C protein in tumor tissues (51% and 57%) was significantly higher than that in adjacent tissues of cancer (9% and 12%) and normal tissues (5% and 6%) (chi2 = 34.6, 38.8, 26.7, 28.6; P < 0.01); It was shown that HPSE and VEGF-C protein expression did significantly not correlate with the type (chi2 = 0.39, 0.41, P > 0.05) and grade of the tumor (chi2 = 0.45, 0.04, P > 0.05); but it correlated with the clinical stage (chi2 = 26.6, 20.1; P < 0.01) and survival time of the patients (chi2 = 21.5, 22.2; P < 0.01). CONCLUSIONS: The results suggest that there be overexpression of HPSE and VEGF-C protein in lung cancer tissues, and which perhaps participate in regulation of tumorigenesis, progression in lung cancer. The expressions of HPSE and VEGF-C protein are used as an useful marker of the biological behavior of lung cancer and as an independent prognosis factor for the patients with lung cancer.